rowid,run.accession,experiment.accession,sample.accession,study.accession,bioproject,study.title,study.alias,study.type,study.abstract,study.attributes,study.PMIDs,sample.description,sample.title,sample.alias,sample.centername,sample.attributes,GEOsample.title,GEOsample.dataprocessing,GEOsample.source,GEOsample.treatmentprotocol,GEOsample.extractprotocol,GEOsample.growthprotocol,GEOsample.characteristics,GEOsample.accession,experiment.title,experiment.alias,experiment.library_name,experiment.design_description,experiment.library_construction_protocol,experiment.attributes,experiment.library_strategy,experiment.library_source,experiment.library_selection,experiment.library_layout,experiment.platform,experiment.instrument_model,experiment.spot_descriptor,experiment.study_ref,run.title,run.attributes,run.filename,run.semantic_name,run.total_bases,run.total_spots,run.alias,run.read_lengths,run.base_counts,run.r1_length,run.r2_length,run.r3_length,run.r4_length,run.Acount,run.Ccount,run.Gcount,run.Tcount,run.Ncount,run.experiment,run.pool_member,submission.accession,submission.srasource,submission.bioprojectsource,seqdetective.n_mates,seqdetective.mapping_rate.mate1,seqdetective.mapping_rate.mate2,seqdetective.nofeature_rate.mate1,seqdetective.nofeature_rate.mate2,seqdetective.sparsity.mate1,seqdetective.sparsity.mate2,seqdetective.pos_strand_rate.mate1,seqdetective.pos_strand_rate.mate2,seqdetective.readlen.mate1,seqdetective.readlen.mate2,seqdetective.judgement.mate1,seqdetective.judgement.mate2,seqdetective.judgement.reason,platform_family,instrument_generation,read_bias,selection_class,prep_kit,sc_or_bulk,tech_class,technology,tech_variant,submission.bioprojectsource.country,earliest_date,devstage_curation,devstage_curation_coarse,tissue_curation,tissue_curation_coarse 9874,ERR4132490,ERX4099806,ERS4552001,ERP121652,PRJEB38247,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,E-MTAB-9054,Transcriptome Analysis,Endocrine disruption can trigger far reaching effects on environmental populations justifying a refusal of market approval for chemicals with ED properties. Ecotoxicogenomic screening was performed to identify molecular fingerprints for endocrine disrupting chemicals affecting the thyroid system in zebrafish Danio rerio embryos as aquatic vertebrate model and alternative to animal testing. 6 Propyl 2 thiouracil 6PTU CAS: 51 52 5 was tested as a model substance for anti thyroidal activity. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of 6PTU for xxx hours under semi static conditions. Each test comprised of a low exposure LE high exposure HE and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked from each sample group and pooled for RNA and protein extraction with NucleoSpin© RNA/Protein kit Macherey Nagel. RNA quality was assessed via a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing 30 million reads per sample. Initial BCL files were demultiplexed to fastq files via bcl2fastq. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library mapped read tables were then merged to a single count matrix. Using this matrix as input read counts were normalized with DESeq2 for differential gene expression analysis.,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. All samples were extracted on the 09.12.2019. CAS: 51 52 5 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. 6 PTU 6 propyl 2 sulfanylidene 1H pyrimidin 4 one CAS number 51 52 5 99% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,R904,SAMEA6824372,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany",ENA FIRST PUBLIC:2020 07 31T04:03:57Z|ENA LAST UPDATE:2020 05 11T17:12:34Z|External Id:SAMEA6824372|INSDC center name:Fraunhofer Institute for Molecular Biology and Applied Ecology Applied Ecology and Bioresources Division Schmallenberg Germany Institute of Ecology Evolution and Diversity Goethe University Frankfurt Frankfurt am Main Germany|INSDC first public:2020 07 31T04:03:57Z|INSDC last update:2020 05 11T17:12:34Z|INSDC status:public|Submitter Id:E MTAB 9054:R904|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|growth condition:low exposure|individual:mixed pool of 10 fish|organism part:whole organism|rin values:10|sample name:E MTAB 9054:R904|scientific name:Danio rerio|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,E MTAB 9054:R904 s,R904 s,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. All samples were extracted on the 09.12.2019. CAS: 51 52 5 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. 6 PTU 6 propyl 2 sulfanylidene 1H pyrimidin 4 one CAS number 51 52 5 99% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,Experimental Factor: compound:6 propyl 2 thiouracil|Experimental Factor: dose:0.001,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP121652,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 11,R904_sr.fastq.gz,fastq,1605165390.0,31794192.0,E MTAB 9054:R904,0:50.49 1:0,A:412564740;C:388045945;G:374953342;T:427657902;N:1943461,50,0,,,412564740,388045945,374953342,427657902,1943461,ERX4099806,ERS4552001,ERA2597157,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive","Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive",1,0.93893,,0.09893,,0.64646,,0.47809,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2020-05-11,Larval,Larval,Whole Organism,All anatomical structures 9875,ERR4132489,ERX4099805,ERS4552000,ERP121652,PRJEB38247,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,E-MTAB-9054,Transcriptome Analysis,Endocrine disruption can trigger far reaching effects on environmental populations justifying a refusal of market approval for chemicals with ED properties. Ecotoxicogenomic screening was performed to identify molecular fingerprints for endocrine disrupting chemicals affecting the thyroid system in zebrafish Danio rerio embryos as aquatic vertebrate model and alternative to animal testing. 6 Propyl 2 thiouracil 6PTU CAS: 51 52 5 was tested as a model substance for anti thyroidal activity. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of 6PTU for xxx hours under semi static conditions. Each test comprised of a low exposure LE high exposure HE and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked from each sample group and pooled for RNA and protein extraction with NucleoSpin© RNA/Protein kit Macherey Nagel. RNA quality was assessed via a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing 30 million reads per sample. Initial BCL files were demultiplexed to fastq files via bcl2fastq. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library mapped read tables were then merged to a single count matrix. Using this matrix as input read counts were normalized with DESeq2 for differential gene expression analysis.,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. All samples were extracted on the 09.12.2019. CAS: 51 52 5 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. 6 PTU 6 propyl 2 sulfanylidene 1H pyrimidin 4 one CAS number 51 52 5 99% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,R900,SAMEA6824371,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany",ENA FIRST PUBLIC:2020 07 31T04:03:57Z|ENA LAST UPDATE:2020 05 11T17:12:34Z|External Id:SAMEA6824371|INSDC center name:Fraunhofer Institute for Molecular Biology and Applied Ecology Applied Ecology and Bioresources Division Schmallenberg Germany Institute of Ecology Evolution and Diversity Goethe University Frankfurt Frankfurt am Main Germany|INSDC first public:2020 07 31T04:03:57Z|INSDC last update:2020 05 11T17:12:34Z|INSDC status:public|Submitter Id:E MTAB 9054:R900|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|growth condition:low exposure|individual:mixed pool of 10 fish|organism part:whole organism|rin values:9.6|sample name:E MTAB 9054:R900|scientific name:Danio rerio|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,E MTAB 9054:R900 s,R900 s,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. All samples were extracted on the 09.12.2019. CAS: 51 52 5 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. 6 PTU 6 propyl 2 sulfanylidene 1H pyrimidin 4 one CAS number 51 52 5 99% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,Experimental Factor: compound:6 propyl 2 thiouracil|Experimental Factor: dose:0.001,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP121652,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 11,R900_sr.fastq.gz,fastq,1597198644.0,31638370.0,E MTAB 9054:R900,0:50.48 1:0,A:410085529;C:386591071;G:373950085;T:424276623;N:2295336,50,0,,,410085529,386591071,373950085,424276623,2295336,ERX4099805,ERS4552000,ERA2597157,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive","Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive",1,0.93876,,0.09801,,0.64889,,0.48057,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2020-05-11,Larval,Larval,Whole Organism,All anatomical structures 9876,ERR4132488,ERX4099804,ERS4551999,ERP121652,PRJEB38247,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,E-MTAB-9054,Transcriptome Analysis,Endocrine disruption can trigger far reaching effects on environmental populations justifying a refusal of market approval for chemicals with ED properties. Ecotoxicogenomic screening was performed to identify molecular fingerprints for endocrine disrupting chemicals affecting the thyroid system in zebrafish Danio rerio embryos as aquatic vertebrate model and alternative to animal testing. 6 Propyl 2 thiouracil 6PTU CAS: 51 52 5 was tested as a model substance for anti thyroidal activity. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of 6PTU for xxx hours under semi static conditions. Each test comprised of a low exposure LE high exposure HE and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked from each sample group and pooled for RNA and protein extraction with NucleoSpin© RNA/Protein kit Macherey Nagel. RNA quality was assessed via a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing 30 million reads per sample. Initial BCL files were demultiplexed to fastq files via bcl2fastq. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library mapped read tables were then merged to a single count matrix. Using this matrix as input read counts were normalized with DESeq2 for differential gene expression analysis.,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. All samples were extracted on the 09.12.2019. CAS: 51 52 5 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. 6 PTU 6 propyl 2 sulfanylidene 1H pyrimidin 4 one CAS number 51 52 5 99% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,R896,SAMEA6824370,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany",ENA FIRST PUBLIC:2020 07 31T04:03:57Z|ENA LAST UPDATE:2020 05 11T17:12:34Z|External Id:SAMEA6824370|INSDC center name:Fraunhofer Institute for Molecular Biology and Applied Ecology Applied Ecology and Bioresources Division Schmallenberg Germany Institute of Ecology Evolution and Diversity Goethe University Frankfurt Frankfurt am Main Germany|INSDC first public:2020 07 31T04:03:57Z|INSDC last update:2020 05 11T17:12:34Z|INSDC status:public|Submitter Id:E MTAB 9054:R896|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|growth condition:low exposure|individual:mixed pool of 10 fish|organism part:whole organism|rin values:10|sample name:E MTAB 9054:R896|scientific name:Danio rerio|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,E MTAB 9054:R896 s,R896 s,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. All samples were extracted on the 09.12.2019. CAS: 51 52 5 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. 6 PTU 6 propyl 2 sulfanylidene 1H pyrimidin 4 one CAS number 51 52 5 99% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,Experimental Factor: compound:6 propyl 2 thiouracil|Experimental Factor: dose:0.001,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP121652,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 11,R896_sr.fastq.gz,fastq,1946340744.0,38553341.0,E MTAB 9054:R896,0:50.48 1:0,A:497881336;C:472722708;G:455800573;T:517246014;N:2690113,50,0,,,497881336,472722708,455800573,517246014,2690113,ERX4099804,ERS4551999,ERA2597157,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive","Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive",1,0.93904,,0.10115,,0.64885,,0.47903,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2020-05-11,Larval,Larval,Whole Organism,All anatomical structures 9877,ERR4132487,ERX4099803,ERS4551998,ERP121652,PRJEB38247,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,E-MTAB-9054,Transcriptome Analysis,Endocrine disruption can trigger far reaching effects on environmental populations justifying a refusal of market approval for chemicals with ED properties. Ecotoxicogenomic screening was performed to identify molecular fingerprints for endocrine disrupting chemicals affecting the thyroid system in zebrafish Danio rerio embryos as aquatic vertebrate model and alternative to animal testing. 6 Propyl 2 thiouracil 6PTU CAS: 51 52 5 was tested as a model substance for anti thyroidal activity. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of 6PTU for xxx hours under semi static conditions. Each test comprised of a low exposure LE high exposure HE and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked from each sample group and pooled for RNA and protein extraction with NucleoSpin© RNA/Protein kit Macherey Nagel. RNA quality was assessed via a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing 30 million reads per sample. Initial BCL files were demultiplexed to fastq files via bcl2fastq. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library mapped read tables were then merged to a single count matrix. Using this matrix as input read counts were normalized with DESeq2 for differential gene expression analysis.,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. All samples were extracted on the 09.12.2019. CAS: 51 52 5 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. 6 PTU 6 propyl 2 sulfanylidene 1H pyrimidin 4 one CAS number 51 52 5 99% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,R906,SAMEA6824369,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany",ENA FIRST PUBLIC:2020 07 31T04:03:57Z|ENA LAST UPDATE:2020 05 11T17:12:34Z|External Id:SAMEA6824369|INSDC center name:Fraunhofer Institute for Molecular Biology and Applied Ecology Applied Ecology and Bioresources Division Schmallenberg Germany Institute of Ecology Evolution and Diversity Goethe University Frankfurt Frankfurt am Main Germany|INSDC first public:2020 07 31T04:03:57Z|INSDC last update:2020 05 11T17:12:34Z|INSDC status:public|Submitter Id:E MTAB 9054:R906|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|growth condition:high exposure|individual:mixed pool of 10 fish|organism part:whole organism|rin values:9.8|sample name:E MTAB 9054:R906|scientific name:Danio rerio|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,E MTAB 9054:R906 s,R906 s,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. All samples were extracted on the 09.12.2019. CAS: 51 52 5 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. 6 PTU 6 propyl 2 sulfanylidene 1H pyrimidin 4 one CAS number 51 52 5 99% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,Experimental Factor: compound:6 propyl 2 thiouracil|Experimental Factor: dose:0.1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP121652,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 11,R906_sr.fastq.gz,fastq,2341343751.0,46379726.0,E MTAB 9054:R906,0:50.48 1:0,A:597650493;C:569437401;G:551299383;T:619558551;N:3397923,50,0,,,597650493,569437401,551299383,619558551,3397923,ERX4099803,ERS4551998,ERA2597157,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive","Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive",1,0.94175,,0.09795,,0.65153,,0.47478,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2020-05-11,Larval,Larval,Whole Organism,All anatomical structures 9878,ERR4132486,ERX4099802,ERS4551997,ERP121652,PRJEB38247,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,E-MTAB-9054,Transcriptome Analysis,Endocrine disruption can trigger far reaching effects on environmental populations justifying a refusal of market approval for chemicals with ED properties. Ecotoxicogenomic screening was performed to identify molecular fingerprints for endocrine disrupting chemicals affecting the thyroid system in zebrafish Danio rerio embryos as aquatic vertebrate model and alternative to animal testing. 6 Propyl 2 thiouracil 6PTU CAS: 51 52 5 was tested as a model substance for anti thyroidal activity. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of 6PTU for xxx hours under semi static conditions. Each test comprised of a low exposure LE high exposure HE and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked from each sample group and pooled for RNA and protein extraction with NucleoSpin© RNA/Protein kit Macherey Nagel. RNA quality was assessed via a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing 30 million reads per sample. Initial BCL files were demultiplexed to fastq files via bcl2fastq. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library mapped read tables were then merged to a single count matrix. Using this matrix as input read counts were normalized with DESeq2 for differential gene expression analysis.,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. All samples were extracted on the 09.12.2019. CAS: 51 52 5 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. 6 PTU 6 propyl 2 sulfanylidene 1H pyrimidin 4 one CAS number 51 52 5 99% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,R902,SAMEA6824368,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany",ENA FIRST PUBLIC:2020 07 31T04:03:57Z|ENA LAST UPDATE:2020 05 11T17:12:34Z|External Id:SAMEA6824368|INSDC center name:Fraunhofer Institute for Molecular Biology and Applied Ecology Applied Ecology and Bioresources Division Schmallenberg Germany Institute of Ecology Evolution and Diversity Goethe University Frankfurt Frankfurt am Main Germany|INSDC first public:2020 07 31T04:03:57Z|INSDC last update:2020 05 11T17:12:34Z|INSDC status:public|Submitter Id:E MTAB 9054:R902|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|growth condition:high exposure|individual:mixed pool of 10 fish|organism part:whole organism|rin values:10|sample name:E MTAB 9054:R902|scientific name:Danio rerio|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,E MTAB 9054:R902 s,R902 s,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. All samples were extracted on the 09.12.2019. CAS: 51 52 5 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. 6 PTU 6 propyl 2 sulfanylidene 1H pyrimidin 4 one CAS number 51 52 5 99% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,Experimental Factor: compound:6 propyl 2 thiouracil|Experimental Factor: dose:0.1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP121652,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 11,R902_sr.fastq.gz,fastq,1632244657.0,32331034.0,E MTAB 9054:R902,0:50.49 1:0,A:413710466;C:400183665;G:387130890;T:428954946;N:2264690,50,0,,,413710466,400183665,387130890,428954946,2264690,ERX4099802,ERS4551997,ERA2597157,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive","Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive",1,0.94282,,0.09186,,0.65192,,0.48446,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2020-05-11,Larval,Larval,Whole Organism,All anatomical structures 9879,ERR4132485,ERX4099801,ERS4551996,ERP121652,PRJEB38247,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,E-MTAB-9054,Transcriptome Analysis,Endocrine disruption can trigger far reaching effects on environmental populations justifying a refusal of market approval for chemicals with ED properties. Ecotoxicogenomic screening was performed to identify molecular fingerprints for endocrine disrupting chemicals affecting the thyroid system in zebrafish Danio rerio embryos as aquatic vertebrate model and alternative to animal testing. 6 Propyl 2 thiouracil 6PTU CAS: 51 52 5 was tested as a model substance for anti thyroidal activity. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of 6PTU for xxx hours under semi static conditions. Each test comprised of a low exposure LE high exposure HE and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked from each sample group and pooled for RNA and protein extraction with NucleoSpin© RNA/Protein kit Macherey Nagel. RNA quality was assessed via a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing 30 million reads per sample. Initial BCL files were demultiplexed to fastq files via bcl2fastq. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library mapped read tables were then merged to a single count matrix. Using this matrix as input read counts were normalized with DESeq2 for differential gene expression analysis.,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. All samples were extracted on the 09.12.2019. CAS: 51 52 5 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. 6 PTU 6 propyl 2 sulfanylidene 1H pyrimidin 4 one CAS number 51 52 5 99% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,R898,SAMEA6824367,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany",ENA FIRST PUBLIC:2020 07 31T04:03:57Z|ENA LAST UPDATE:2020 05 11T17:12:34Z|External Id:SAMEA6824367|INSDC center name:Fraunhofer Institute for Molecular Biology and Applied Ecology Applied Ecology and Bioresources Division Schmallenberg Germany Institute of Ecology Evolution and Diversity Goethe University Frankfurt Frankfurt am Main Germany|INSDC first public:2020 07 31T04:03:57Z|INSDC last update:2020 05 11T17:12:34Z|INSDC status:public|Submitter Id:E MTAB 9054:R898|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|growth condition:high exposure|individual:mixed pool of 10 fish|organism part:whole organism|rin values:9.8|sample name:E MTAB 9054:R898|scientific name:Danio rerio|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,E MTAB 9054:R898 s,R898 s,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. All samples were extracted on the 09.12.2019. CAS: 51 52 5 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. 6 PTU 6 propyl 2 sulfanylidene 1H pyrimidin 4 one CAS number 51 52 5 99% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,Experimental Factor: compound:6 propyl 2 thiouracil|Experimental Factor: dose:0.1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP121652,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 11,R898_sr.fastq.gz,fastq,1973588979.0,39097363.0,E MTAB 9054:R898,0:50.48 1:0,A:501700006;C:481059078;G:466931881;T:520778140;N:3119874,50,0,,,501700006,481059078,466931881,520778140,3119874,ERX4099801,ERS4551996,ERA2597157,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive","Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive",1,0.94133,,0.09578,,0.651,,0.474,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2020-05-11,Larval,Larval,Whole Organism,All anatomical structures 9880,ERR4132484,ERX4099800,ERS4551995,ERP121652,PRJEB38247,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,E-MTAB-9054,Transcriptome Analysis,Endocrine disruption can trigger far reaching effects on environmental populations justifying a refusal of market approval for chemicals with ED properties. Ecotoxicogenomic screening was performed to identify molecular fingerprints for endocrine disrupting chemicals affecting the thyroid system in zebrafish Danio rerio embryos as aquatic vertebrate model and alternative to animal testing. 6 Propyl 2 thiouracil 6PTU CAS: 51 52 5 was tested as a model substance for anti thyroidal activity. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of 6PTU for xxx hours under semi static conditions. Each test comprised of a low exposure LE high exposure HE and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked from each sample group and pooled for RNA and protein extraction with NucleoSpin© RNA/Protein kit Macherey Nagel. RNA quality was assessed via a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing 30 million reads per sample. Initial BCL files were demultiplexed to fastq files via bcl2fastq. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library mapped read tables were then merged to a single count matrix. Using this matrix as input read counts were normalized with DESeq2 for differential gene expression analysis.,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. All samples were extracted on the 09.12.2019. CAS: 51 52 5 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. 6 PTU 6 propyl 2 sulfanylidene 1H pyrimidin 4 one CAS number 51 52 5 99% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,R903,SAMEA6824366,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany",ENA FIRST PUBLIC:2020 07 31T04:03:57Z|ENA LAST UPDATE:2020 05 11T17:12:34Z|External Id:SAMEA6824366|INSDC center name:Fraunhofer Institute for Molecular Biology and Applied Ecology Applied Ecology and Bioresources Division Schmallenberg Germany Institute of Ecology Evolution and Diversity Goethe University Frankfurt Frankfurt am Main Germany|INSDC first public:2020 07 31T04:03:57Z|INSDC last update:2020 05 11T17:12:34Z|INSDC status:public|Submitter Id:E MTAB 9054:R903|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|growth condition:normal|individual:mixed pool of 10 fish|organism part:whole organism|rin values:9.8|sample name:E MTAB 9054:R903|scientific name:Danio rerio|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,E MTAB 9054:R903 s,R903 s,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. All samples were extracted on the 09.12.2019. CAS: 51 52 5 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. 6 PTU 6 propyl 2 sulfanylidene 1H pyrimidin 4 one CAS number 51 52 5 99% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP121652,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 11,R903_sr.fastq.gz,fastq,1978975800.0,39199481.0,E MTAB 9054:R903,0:50.48 1:0,A:502614523;C:482986303;G:468168184;T:522608062;N:2598728,50,0,,,502614523,482986303,468168184,522608062,2598728,ERX4099800,ERS4551995,ERA2597157,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive","Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive",1,0.9404,,0.09216,,0.64607,,0.47417,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2020-05-11,Larval,Larval,Whole Organism,All anatomical structures 9881,ERR4132483,ERX4099799,ERS4551994,ERP121652,PRJEB38247,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,E-MTAB-9054,Transcriptome Analysis,Endocrine disruption can trigger far reaching effects on environmental populations justifying a refusal of market approval for chemicals with ED properties. Ecotoxicogenomic screening was performed to identify molecular fingerprints for endocrine disrupting chemicals affecting the thyroid system in zebrafish Danio rerio embryos as aquatic vertebrate model and alternative to animal testing. 6 Propyl 2 thiouracil 6PTU CAS: 51 52 5 was tested as a model substance for anti thyroidal activity. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of 6PTU for xxx hours under semi static conditions. Each test comprised of a low exposure LE high exposure HE and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked from each sample group and pooled for RNA and protein extraction with NucleoSpin© RNA/Protein kit Macherey Nagel. RNA quality was assessed via a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing 30 million reads per sample. Initial BCL files were demultiplexed to fastq files via bcl2fastq. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library mapped read tables were then merged to a single count matrix. Using this matrix as input read counts were normalized with DESeq2 for differential gene expression analysis.,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. All samples were extracted on the 09.12.2019. CAS: 51 52 5 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. 6 PTU 6 propyl 2 sulfanylidene 1H pyrimidin 4 one CAS number 51 52 5 99% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,R899,SAMEA6824365,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany",ENA FIRST PUBLIC:2020 07 31T04:03:57Z|ENA LAST UPDATE:2020 05 11T17:12:34Z|External Id:SAMEA6824365|INSDC center name:Fraunhofer Institute for Molecular Biology and Applied Ecology Applied Ecology and Bioresources Division Schmallenberg Germany Institute of Ecology Evolution and Diversity Goethe University Frankfurt Frankfurt am Main Germany|INSDC first public:2020 07 31T04:03:57Z|INSDC last update:2020 05 11T17:12:34Z|INSDC status:public|Submitter Id:E MTAB 9054:R899|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|growth condition:normal|individual:mixed pool of 10 fish|organism part:whole organism|rin values:10|sample name:E MTAB 9054:R899|scientific name:Danio rerio|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,E MTAB 9054:R899 s,R899 s,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. All samples were extracted on the 09.12.2019. CAS: 51 52 5 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. 6 PTU 6 propyl 2 sulfanylidene 1H pyrimidin 4 one CAS number 51 52 5 99% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP121652,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 11,R899_sr.fastq.gz,fastq,1556233879.0,30835141.0,E MTAB 9054:R899,0:50.47 1:0,A:394448696;C:380284673;G:367546543;T:410802983;N:3150984,50,0,,,394448696,380284673,367546543,410802983,3150984,ERX4099799,ERS4551994,ERA2597157,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive","Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive",1,0.93845,,0.09509,,0.64926,,0.48436,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2020-05-11,Larval,Larval,Whole Organism,All anatomical structures 9882,ERR4132482,ERX4099798,ERS4551993,ERP121652,PRJEB38247,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,E-MTAB-9054,Transcriptome Analysis,Endocrine disruption can trigger far reaching effects on environmental populations justifying a refusal of market approval for chemicals with ED properties. Ecotoxicogenomic screening was performed to identify molecular fingerprints for endocrine disrupting chemicals affecting the thyroid system in zebrafish Danio rerio embryos as aquatic vertebrate model and alternative to animal testing. 6 Propyl 2 thiouracil 6PTU CAS: 51 52 5 was tested as a model substance for anti thyroidal activity. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of 6PTU for xxx hours under semi static conditions. Each test comprised of a low exposure LE high exposure HE and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked from each sample group and pooled for RNA and protein extraction with NucleoSpin© RNA/Protein kit Macherey Nagel. RNA quality was assessed via a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing 30 million reads per sample. Initial BCL files were demultiplexed to fastq files via bcl2fastq. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library mapped read tables were then merged to a single count matrix. Using this matrix as input read counts were normalized with DESeq2 for differential gene expression analysis.,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. All samples were extracted on the 09.12.2019. CAS: 51 52 5 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. 6 PTU 6 propyl 2 sulfanylidene 1H pyrimidin 4 one CAS number 51 52 5 99% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,R895,SAMEA6824364,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany",ENA FIRST PUBLIC:2020 07 31T04:03:57Z|ENA LAST UPDATE:2020 05 11T17:12:34Z|External Id:SAMEA6824364|INSDC center name:Fraunhofer Institute for Molecular Biology and Applied Ecology Applied Ecology and Bioresources Division Schmallenberg Germany Institute of Ecology Evolution and Diversity Goethe University Frankfurt Frankfurt am Main Germany|INSDC first public:2020 07 31T04:03:57Z|INSDC last update:2020 05 11T17:12:34Z|INSDC status:public|Submitter Id:E MTAB 9054:R895|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|growth condition:normal|individual:mixed pool of 10 fish|organism part:whole organism|rin values:10|sample name:E MTAB 9054:R895|scientific name:Danio rerio|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,E MTAB 9054:R895 s,R895 s,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. All samples were extracted on the 09.12.2019. CAS: 51 52 5 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. 6 PTU 6 propyl 2 sulfanylidene 1H pyrimidin 4 one CAS number 51 52 5 99% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP121652,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of 6 Propyl 2 thiouracil 6 PTU below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 11,R895_sr.fastq.gz,fastq,1098959757.0,21771883.0,E MTAB 9054:R895,0:50.48 1:0,A:281414032;C:266978094;G:257304686;T:291591340;N:1671605,50,0,,,281414032,266978094,257304686,291591340,1671605,ERX4099798,ERS4551993,ERA2597157,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive","Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive",1,0.93996,,0.09776,,0.64989,,0.47968,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2020-05-11,Larval,Larval,Whole Organism,All anatomical structures 9883,ERR4140034,ERX4107347,ERS4556110,ERP121678,PRJEB38271,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,E-MTAB-9056,Transcriptome Analysis,The aim of this sequencing experiment was to screen for ecotoxicogenomic fingerprints for endocrine disrupting chemicals affecting the thyroid system in zebrafish Danio rerio embryos as aquatic vertebrate model and alternative to animal testing. Triiodothyronine T3 CAS: 6893 02 3 was tested as a model substance for thyroidal inducing activity. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of T3 for xxx hours under semi static conditions. Each test comprised of a low exposure LE high exposure HE and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 12,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. T3 3 three prime 5 Triiodo L thyronine CAS number 6893 02 3 95% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,R86,SAMEA6828487,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany",ENA FIRST PUBLIC:2020 07 31T04:03:57Z|ENA LAST UPDATE:2020 05 12T15:52:19Z|External Id:SAMEA6828487|INSDC center name:Fraunhofer Institute for Molecular Biology and Applied Ecology Applied Ecology and Bioresources Division Schmallenberg Germany Institute of Ecology Evolution and Diversity Goethe University Frankfurt Frankfurt am Main Germany|INSDC first public:2020 07 31T04:03:57Z|INSDC last update:2020 05 12T15:52:19Z|INSDC status:public|Submitter Id:E MTAB 9056:R86|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|growth condition:low exposure|individual:pool of 10 fish|organism part:whole organism|sample name:E MTAB 9056:R86|scientific name:Danio rerio|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyro9 T3 below acute toxicity levels against untreated control groups,E MTAB 9056:R86 s,R86 s,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. T3 3 three prime 5 Triiodo L thyronine CAS number 6893 02 3 95% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,Experimental Factor: compound:3 3 prime 5 triiodo L thyro9|Experimental Factor: dose:3.3E 06,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP121678,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 13,R86_sr.fastq.gz,fastq,1327826966.0,26294265.0,E MTAB 9056:R86,0:50.50 1:0,A:340934934;C:322947435;G:309801278;T:353159102;N:984217,50,0,,,340934934,322947435,309801278,353159102,984217,ERX4107347,ERS4556110,ERA2597448,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive","Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive",1,0.9393,,0.1031,,0.65683,,0.46848,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2020-05-12,Larval,Larval,Whole Organism,All anatomical structures 9884,ERR4140033,ERX4107346,ERS4556109,ERP121678,PRJEB38271,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,E-MTAB-9056,Transcriptome Analysis,The aim of this sequencing experiment was to screen for ecotoxicogenomic fingerprints for endocrine disrupting chemicals affecting the thyroid system in zebrafish Danio rerio embryos as aquatic vertebrate model and alternative to animal testing. Triiodothyronine T3 CAS: 6893 02 3 was tested as a model substance for thyroidal inducing activity. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of T3 for xxx hours under semi static conditions. Each test comprised of a low exposure LE high exposure HE and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 12,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. T3 3 three prime 5 Triiodo L thyronine CAS number 6893 02 3 95% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,R164,SAMEA6828486,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany",ENA FIRST PUBLIC:2020 07 31T04:03:57Z|ENA LAST UPDATE:2020 05 12T15:52:19Z|External Id:SAMEA6828486|INSDC center name:Fraunhofer Institute for Molecular Biology and Applied Ecology Applied Ecology and Bioresources Division Schmallenberg Germany Institute of Ecology Evolution and Diversity Goethe University Frankfurt Frankfurt am Main Germany|INSDC first public:2020 07 31T04:03:57Z|INSDC last update:2020 05 12T15:52:19Z|INSDC status:public|Submitter Id:E MTAB 9056:R164|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|growth condition:low exposure|individual:pool of 10 fish|organism part:whole organism|sample name:E MTAB 9056:R164|scientific name:Danio rerio|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyro9 T3 below acute toxicity levels against untreated control groups,E MTAB 9056:R164 s,R164 s,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. T3 3 three prime 5 Triiodo L thyronine CAS number 6893 02 3 95% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,Experimental Factor: compound:3 3 prime 5 triiodo L thyro9|Experimental Factor: dose:3.3E 06,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP121678,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 13,R164_sr.fastq.gz,fastq,1513567439.0,29975869.0,E MTAB 9056:R164,0:50.49 1:0,A:388109813;C:368964600;G:352756505;T:402261160;N:1475361,50,0,,,388109813,368964600,352756505,402261160,1475361,ERX4107346,ERS4556109,ERA2597448,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive","Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive",1,0.93992,,0.10282,,0.65437,,0.47367,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2020-05-12,Larval,Larval,Whole Organism,All anatomical structures 9885,ERR4140032,ERX4107345,ERS4556108,ERP121678,PRJEB38271,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,E-MTAB-9056,Transcriptome Analysis,The aim of this sequencing experiment was to screen for ecotoxicogenomic fingerprints for endocrine disrupting chemicals affecting the thyroid system in zebrafish Danio rerio embryos as aquatic vertebrate model and alternative to animal testing. Triiodothyronine T3 CAS: 6893 02 3 was tested as a model substance for thyroidal inducing activity. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of T3 for xxx hours under semi static conditions. Each test comprised of a low exposure LE high exposure HE and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 12,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. T3 3 three prime 5 Triiodo L thyronine CAS number 6893 02 3 95% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,R158,SAMEA6828485,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany",ENA FIRST PUBLIC:2020 07 31T04:03:57Z|ENA LAST UPDATE:2020 05 12T15:52:19Z|External Id:SAMEA6828485|INSDC center name:Fraunhofer Institute for Molecular Biology and Applied Ecology Applied Ecology and Bioresources Division Schmallenberg Germany Institute of Ecology Evolution and Diversity Goethe University Frankfurt Frankfurt am Main Germany|INSDC first public:2020 07 31T04:03:57Z|INSDC last update:2020 05 12T15:52:19Z|INSDC status:public|Submitter Id:E MTAB 9056:R158|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|growth condition:low exposure|individual:pool of 10 fish|organism part:whole organism|sample name:E MTAB 9056:R158|scientific name:Danio rerio|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyro9 T3 below acute toxicity levels against untreated control groups,E MTAB 9056:R158 s,R158 s,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. T3 3 three prime 5 Triiodo L thyronine CAS number 6893 02 3 95% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,Experimental Factor: compound:3 3 prime 5 triiodo L thyro9|Experimental Factor: dose:3.3E 06,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP121678,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 13,R158_sr.fastq.gz,fastq,1512642153.0,29962837.0,E MTAB 9056:R158,0:50.48 1:0,A:389585690;C:365364747;G:351299223;T:404327286;N:2065207,50,0,,,389585690,365364747,351299223,404327286,2065207,ERX4107345,ERS4556108,ERA2597448,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive","Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive",1,0.93757,,0.10553,,0.65557,,0.47147,,49,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2020-05-12,Larval,Larval,Whole Organism,All anatomical structures 9886,ERR4140031,ERX4107344,ERS4556107,ERP121678,PRJEB38271,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,E-MTAB-9056,Transcriptome Analysis,The aim of this sequencing experiment was to screen for ecotoxicogenomic fingerprints for endocrine disrupting chemicals affecting the thyroid system in zebrafish Danio rerio embryos as aquatic vertebrate model and alternative to animal testing. Triiodothyronine T3 CAS: 6893 02 3 was tested as a model substance for thyroidal inducing activity. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of T3 for xxx hours under semi static conditions. Each test comprised of a low exposure LE high exposure HE and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 12,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. T3 3 three prime 5 Triiodo L thyronine CAS number 6893 02 3 95% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,R89,SAMEA6828484,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany",ENA FIRST PUBLIC:2020 07 31T04:03:57Z|ENA LAST UPDATE:2020 05 12T15:52:19Z|External Id:SAMEA6828484|INSDC center name:Fraunhofer Institute for Molecular Biology and Applied Ecology Applied Ecology and Bioresources Division Schmallenberg Germany Institute of Ecology Evolution and Diversity Goethe University Frankfurt Frankfurt am Main Germany|INSDC first public:2020 07 31T04:03:57Z|INSDC last update:2020 05 12T15:52:19Z|INSDC status:public|Submitter Id:E MTAB 9056:R89|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|growth condition:high exposure|individual:pool of 10 fish|organism part:whole organism|sample name:E MTAB 9056:R89|scientific name:Danio rerio|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyro9 T3 below acute toxicity levels against untreated control groups,E MTAB 9056:R89 s,R89 s,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. T3 3 three prime 5 Triiodo L thyronine CAS number 6893 02 3 95% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,Experimental Factor: compound:3 3 prime 5 triiodo L thyro9|Experimental Factor: dose:0.000327,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP121678,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 13,R89_sr.fastq.gz,fastq,1347758243.0,26686154.0,E MTAB 9056:R89,0:50.50 1:0,A:347954623;C:326744319;G:311736057;T:360479809;N:843435,50,0,,,347954623,326744319,311736057,360479809,843435,ERX4107344,ERS4556107,ERA2597448,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive","Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive",1,0.93909,,0.1032,,0.64906,,0.47499,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2020-05-12,Larval,Larval,Whole Organism,All anatomical structures 9887,ERR4140030,ERX4107343,ERS4556106,ERP121678,PRJEB38271,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,E-MTAB-9056,Transcriptome Analysis,The aim of this sequencing experiment was to screen for ecotoxicogenomic fingerprints for endocrine disrupting chemicals affecting the thyroid system in zebrafish Danio rerio embryos as aquatic vertebrate model and alternative to animal testing. Triiodothyronine T3 CAS: 6893 02 3 was tested as a model substance for thyroidal inducing activity. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of T3 for xxx hours under semi static conditions. Each test comprised of a low exposure LE high exposure HE and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 12,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. T3 3 three prime 5 Triiodo L thyronine CAS number 6893 02 3 95% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,R167,SAMEA6828483,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany",ENA FIRST PUBLIC:2020 07 31T04:03:57Z|ENA LAST UPDATE:2020 05 12T15:52:19Z|External Id:SAMEA6828483|INSDC center name:Fraunhofer Institute for Molecular Biology and Applied Ecology Applied Ecology and Bioresources Division Schmallenberg Germany Institute of Ecology Evolution and Diversity Goethe University Frankfurt Frankfurt am Main Germany|INSDC first public:2020 07 31T04:03:57Z|INSDC last update:2020 05 12T15:52:19Z|INSDC status:public|Submitter Id:E MTAB 9056:R167|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|growth condition:high exposure|individual:pool of 10 fish|organism part:whole organism|sample name:E MTAB 9056:R167|scientific name:Danio rerio|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyro9 T3 below acute toxicity levels against untreated control groups,E MTAB 9056:R167 s,R167 s,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. T3 3 three prime 5 Triiodo L thyronine CAS number 6893 02 3 95% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,Experimental Factor: compound:3 3 prime 5 triiodo L thyro9|Experimental Factor: dose:0.000327,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP121678,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 13,R167_sr.fastq.gz,fastq,1422756236.0,28174340.0,E MTAB 9056:R167,0:50.50 1:0,A:366380887;C:345824655;G:330108682;T:379312527;N:1129485,50,0,,,366380887,345824655,330108682,379312527,1129485,ERX4107343,ERS4556106,ERA2597448,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive","Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive",1,0.93724,,0.10506,,0.65001,,0.47599,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2020-05-12,Larval,Larval,Whole Organism,All anatomical structures 9888,ERR4140029,ERX4107342,ERS4556105,ERP121678,PRJEB38271,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,E-MTAB-9056,Transcriptome Analysis,The aim of this sequencing experiment was to screen for ecotoxicogenomic fingerprints for endocrine disrupting chemicals affecting the thyroid system in zebrafish Danio rerio embryos as aquatic vertebrate model and alternative to animal testing. Triiodothyronine T3 CAS: 6893 02 3 was tested as a model substance for thyroidal inducing activity. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of T3 for xxx hours under semi static conditions. Each test comprised of a low exposure LE high exposure HE and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 12,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. T3 3 three prime 5 Triiodo L thyronine CAS number 6893 02 3 95% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,R161,SAMEA6828482,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany",ENA FIRST PUBLIC:2020 07 31T04:03:57Z|ENA LAST UPDATE:2020 05 12T15:52:19Z|External Id:SAMEA6828482|INSDC center name:Fraunhofer Institute for Molecular Biology and Applied Ecology Applied Ecology and Bioresources Division Schmallenberg Germany Institute of Ecology Evolution and Diversity Goethe University Frankfurt Frankfurt am Main Germany|INSDC first public:2020 07 31T04:03:57Z|INSDC last update:2020 05 12T15:52:19Z|INSDC status:public|Submitter Id:E MTAB 9056:R161|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|growth condition:high exposure|individual:pool of 10 fish|organism part:whole organism|sample name:E MTAB 9056:R161|scientific name:Danio rerio|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyro9 T3 below acute toxicity levels against untreated control groups,E MTAB 9056:R161 s,R161 s,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. T3 3 three prime 5 Triiodo L thyronine CAS number 6893 02 3 95% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,Experimental Factor: compound:3 3 prime 5 triiodo L thyro9|Experimental Factor: dose:0.000327,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP121678,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 13,R161_sr.fastq.gz,fastq,1696412529.0,33592555.0,E MTAB 9056:R161,0:50.50 1:0,A:439133160;C:409023953;G:389977077;T:457053986;N:1224353,50,0,,,439133160,409023953,389977077,457053986,1224353,ERX4107342,ERS4556105,ERA2597448,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive","Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive",1,0.93609,,0.1083,,0.6535,,0.46658,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2020-05-12,Larval,Larval,Whole Organism,All anatomical structures 9889,ERR4140028,ERX4107341,ERS4556104,ERP121678,PRJEB38271,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,E-MTAB-9056,Transcriptome Analysis,The aim of this sequencing experiment was to screen for ecotoxicogenomic fingerprints for endocrine disrupting chemicals affecting the thyroid system in zebrafish Danio rerio embryos as aquatic vertebrate model and alternative to animal testing. Triiodothyronine T3 CAS: 6893 02 3 was tested as a model substance for thyroidal inducing activity. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of T3 for xxx hours under semi static conditions. Each test comprised of a low exposure LE high exposure HE and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 12,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. T3 3 three prime 5 Triiodo L thyronine CAS number 6893 02 3 95% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,R85,SAMEA6828481,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany",ENA FIRST PUBLIC:2020 07 31T04:03:57Z|ENA LAST UPDATE:2020 05 12T15:52:19Z|External Id:SAMEA6828481|INSDC center name:Fraunhofer Institute for Molecular Biology and Applied Ecology Applied Ecology and Bioresources Division Schmallenberg Germany Institute of Ecology Evolution and Diversity Goethe University Frankfurt Frankfurt am Main Germany|INSDC first public:2020 07 31T04:03:57Z|INSDC last update:2020 05 12T15:52:19Z|INSDC status:public|Submitter Id:E MTAB 9056:R85|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|growth condition:control|individual:pool of 10 fish|organism part:whole organism|sample name:E MTAB 9056:R85|scientific name:Danio rerio|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyro9 T3 below acute toxicity levels against untreated control groups,E MTAB 9056:R85 s,R85 s,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. T3 3 three prime 5 Triiodo L thyronine CAS number 6893 02 3 95% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,Experimental Factor: compound:n1|Experimental Factor: dose:0,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP121678,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 13,R85_sr.fastq.gz,fastq,1115426084.0,22086160.0,E MTAB 9056:R85,0:50.50 1:0,A:288411880;C:265358022;G:259432943;T:301493661;N:729578,50,0,,,288411880,265358022,259432943,301493661,729578,ERX4107341,ERS4556104,ERA2597448,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive","Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive",1,0.94238,,0.10918,,0.65372,,0.48727,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2020-05-12,Larval,Larval,Whole Organism,All anatomical structures 9890,ERR4140027,ERX4107340,ERS4556103,ERP121678,PRJEB38271,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,E-MTAB-9056,Transcriptome Analysis,The aim of this sequencing experiment was to screen for ecotoxicogenomic fingerprints for endocrine disrupting chemicals affecting the thyroid system in zebrafish Danio rerio embryos as aquatic vertebrate model and alternative to animal testing. Triiodothyronine T3 CAS: 6893 02 3 was tested as a model substance for thyroidal inducing activity. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of T3 for xxx hours under semi static conditions. Each test comprised of a low exposure LE high exposure HE and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 12,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. T3 3 three prime 5 Triiodo L thyronine CAS number 6893 02 3 95% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,R162,SAMEA6828480,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany",ENA FIRST PUBLIC:2020 07 31T04:03:57Z|ENA LAST UPDATE:2020 05 12T15:52:19Z|External Id:SAMEA6828480|INSDC center name:Fraunhofer Institute for Molecular Biology and Applied Ecology Applied Ecology and Bioresources Division Schmallenberg Germany Institute of Ecology Evolution and Diversity Goethe University Frankfurt Frankfurt am Main Germany|INSDC first public:2020 07 31T04:03:57Z|INSDC last update:2020 05 12T15:52:19Z|INSDC status:public|Submitter Id:E MTAB 9056:R162|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|growth condition:control|individual:pool of 10 fish|organism part:whole organism|sample name:E MTAB 9056:R162|scientific name:Danio rerio|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyro9 T3 below acute toxicity levels against untreated control groups,E MTAB 9056:R162 s,R162 s,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. T3 3 three prime 5 Triiodo L thyronine CAS number 6893 02 3 95% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,Experimental Factor: compound:n1|Experimental Factor: dose:0,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP121678,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 13,R162_sr.fastq.gz,fastq,1368462525.0,27098537.0,E MTAB 9056:R162,0:50.50 1:0,A:352965195;C:330975762;G:317956233;T:365598349;N:966986,50,0,,,352965195,330975762,317956233,365598349,966986,ERX4107340,ERS4556103,ERA2597448,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive","Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive",1,0.93788,,0.10812,,0.65482,,0.47962,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2020-05-12,Larval,Larval,Whole Organism,All anatomical structures 9891,ERR4140026,ERX4107339,ERS4556102,ERP121678,PRJEB38271,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,E-MTAB-9056,Transcriptome Analysis,The aim of this sequencing experiment was to screen for ecotoxicogenomic fingerprints for endocrine disrupting chemicals affecting the thyroid system in zebrafish Danio rerio embryos as aquatic vertebrate model and alternative to animal testing. Triiodothyronine T3 CAS: 6893 02 3 was tested as a model substance for thyroidal inducing activity. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of T3 for xxx hours under semi static conditions. Each test comprised of a low exposure LE high exposure HE and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 12,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. T3 3 three prime 5 Triiodo L thyronine CAS number 6893 02 3 95% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,R157,SAMEA6828479,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany",ENA FIRST PUBLIC:2020 07 31T04:03:57Z|ENA LAST UPDATE:2020 05 12T15:52:19Z|External Id:SAMEA6828479|INSDC center name:Fraunhofer Institute for Molecular Biology and Applied Ecology Applied Ecology and Bioresources Division Schmallenberg Germany Institute of Ecology Evolution and Diversity Goethe University Frankfurt Frankfurt am Main Germany|INSDC first public:2020 07 31T04:03:57Z|INSDC last update:2020 05 12T15:52:19Z|INSDC status:public|Submitter Id:E MTAB 9056:R157|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|growth condition:control|individual:pool of 10 fish|organism part:whole organism|sample name:E MTAB 9056:R157|scientific name:Danio rerio|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyro9 T3 below acute toxicity levels against untreated control groups,E MTAB 9056:R157 s,R157 s,RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were picked for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. T3 3 three prime 5 Triiodo L thyronine CAS number 6893 02 3 95% purity was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark while being constantly aerated. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 40 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined through a 2100 Bioanalyzer© Instrument system Agilent G2939BA using RNApico© chips Agilent 5067 1513. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina.,Experimental Factor: compound:n1|Experimental Factor: dose:0,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP121678,Illumina HiSeq 4000 sequencing; RNA Seq of zebrafish embryos 96hpf treated with different concentrations of Triiodothyronine T3 below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2020 09 30|ENA LAST UPDATE:2020 05 13,R157_sr.fastq.gz,fastq,1500167669.0,29707076.0,E MTAB 9056:R157,0:50.50 1:0,A:386622926;C:363330639;G:347819057;T:401278075;N:1116972,50,0,,,386622926,363330639,347819057,401278075,1116972,ERX4107339,ERS4556102,ERA2597448,"Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive","Fraunhofer Institute for Molecular Biology and Applied Ecology, Applied Ecology and Bioresources Division, Schmallenberg, Germany Institute of Ecology, Evolution and Diversity, Goethe University Frankfurt, Frankfurt am Main, Germany|European Nucleotide Archive",1,0.93887,,0.10641,,0.65283,,0.46616,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2020-05-12,Larval,Larval,Whole Organism,All anatomical structures 10068,ERR4844843,ERX4714625,ERS5338302,ERP125162,PRJEB41393,Depletion of Foxk transcription factors causes genome wide transcriptional misregulation and developmental arrest in zebrafish embryos,E-MTAB-9777,Transcriptome Analysis,Foxk proteins are transcriptional regulators implicated in key biological processes such as glycolysis autophagy and cell cycle regulation among others. Here we employ targeted morpholino knockdown to deplete Foxk1 Fokx2 and Foxk2 1 proteins in developing zebrafish embryos. We demonstrate that the loss of Foxk transcription factors causes genome wide transcriptional misregulation characterised by upregulation of autophagy related genes and downregulation of cell cycle regulators. The phenotype is embryonic lethal with the majority of embryos not surviving past 24hpf.,ENA FIRST PUBLIC:2020 12 01|ENA LAST UPDATE:2020 11 17,,Protocols: Embryos were collected 0 hpf and incubated in 1X E3 medium 0.03% NaCl 0.005% CaCl2 0.0013% KCl 99.9557% H2O 0.008% H14MgO11S for 24 hours at 28.5°C Adult wild type AB/Tubingen Danio rerio zebrafish were bred in a 1 male:1 female ratio. 1c embryos were injected with morpholino oligonucleotides targeting foxk1 foxk2 and foxk2 1 transcripts Total RNA was purified from xxx embryos using the RNeasy Mini Kit Qiagen Valencia CA USA according to the manufacturer's instructions. mRNA Seq libraries were generated from total RNA with polyA+ selection of mRNA using the TruSeq RNA Sample Prep Kit v2 Illumina San Diego CA. Strand specific libraries were constructed using a dUTP methodology as described previously [Zhong S. et al. High Throughput Illumina Strand Specific RNA Sequencing Library Preparation. Cold Spring Harb. Protoc. 2011 940 949 2011],wt rep2,SAMEA7579966,Garvan Institute of Medical Research,ENA first public:2020 12 01|ENA last update:2020 11 17|External Id:SAMEA7579966|INSDC center alias:Garvan Institute of Medical Research|INSDC center name:Garvan Institute of Medical Research|INSDC first public:2020 12 01T04:12:08Z|INSDC last update:2020 11 17T14:25:39Z|INSDC status:public|Submitter Id:E MTAB 9777:wt rep2|age:24hpf|broker name:ArrayExpress|common name:zebrafish|developmental stage:embryo stage|organism part:whole organism|sample name:E MTAB 9777:wt rep2|strain:Ab / Tubingen,,,,,,,,,Illumina HiSeq 1500 sequencing; Depletion of Foxk transcription factors causes genome wide transcriptional misregulation and developmental arrest in zebrafish embryos,E MTAB 9777:wt rep2 s,wt rep2 s,Depletion of Foxk transcription factors causes genome wide transcriptional misregulation and developmental arrest in zebrafish embryos,Embryos were collected 0 hpf and incubated in 1X E3 medium 0.03% NaCl 0.005% CaCl2 0.0013% KCl 99.9557% H2O 0.008% H14MgO11S for 24 hours at 28.5°C Adult wild type AB/Tubingen Danio rerio zebrafish were bred in a 1 male:1 female ratio. 1c embryos were injected with morpholino oligonucleotides targeting foxk1 foxk2 and foxk2 1 transcripts Total RNA was purified from xxx embryos using the RNeasy Mini Kit Qiagen Valencia CA USA according to the manufacturer's instructions. mRNA Seq libraries were generated from total RNA with polyA+ selection of mRNA using the TruSeq RNA Sample Prep Kit v2 Illumina San Diego CA. Strand specific libraries were constructed using a dUTP methodology as described previously [Zhong S. et al. High Throughput Illumina Strand Specific RNA Sequencing Library Preparation. Cold Spring Harb. Protoc. 2011 940 949 2011],Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 1500,,ERP125162,Illumina HiSeq 1500 sequencing; Depletion of Foxk transcription factors causes genome wide transcriptional misregulation and developmental arrest in zebrafish embryos,ENA FIRST PUBLIC:2020 12 01|ENA LAST UPDATE:2020 11 17,wt_rep2.fastq.gz,fastq,3456340392.0,34221192.0,E MTAB 9777:wt rep2,0:101 1:0,A:850781866;C:827613239;G:837337477;T:936245388;N:4362422,101,0,,,850781866,827613239,837337477,936245388,4362422,ERX4714625,ERS5338302,ERA3145932,European Nucleotide Archive,European Nucleotide Archive,1,0.94198,,0.05681,,0.69613,,0.46685,,101,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Unknown,2020-11-17,Pharyngula,Embryo,Whole Organism,All anatomical structures 10069,ERR4844842,ERX4714624,ERS5338301,ERP125162,PRJEB41393,Depletion of Foxk transcription factors causes genome wide transcriptional misregulation and developmental arrest in zebrafish embryos,E-MTAB-9777,Transcriptome Analysis,Foxk proteins are transcriptional regulators implicated in key biological processes such as glycolysis autophagy and cell cycle regulation among others. Here we employ targeted morpholino knockdown to deplete Foxk1 Fokx2 and Foxk2 1 proteins in developing zebrafish embryos. We demonstrate that the loss of Foxk transcription factors causes genome wide transcriptional misregulation characterised by upregulation of autophagy related genes and downregulation of cell cycle regulators. The phenotype is embryonic lethal with the majority of embryos not surviving past 24hpf.,ENA FIRST PUBLIC:2020 12 01|ENA LAST UPDATE:2020 11 17,,Protocols: Embryos were collected 0 hpf and incubated in 1X E3 medium 0.03% NaCl 0.005% CaCl2 0.0013% KCl 99.9557% H2O 0.008% H14MgO11S for 24 hours at 28.5°C Adult wild type AB/Tubingen Danio rerio zebrafish were bred in a 1 male:1 female ratio. 1c embryos were injected with morpholino oligonucleotides targeting foxk1 foxk2 and foxk2 1 transcripts Total RNA was purified from xxx embryos using the RNeasy Mini Kit Qiagen Valencia CA USA according to the manufacturer's instructions. mRNA Seq libraries were generated from total RNA with polyA+ selection of mRNA using the TruSeq RNA Sample Prep Kit v2 Illumina San Diego CA. Strand specific libraries were constructed using a dUTP methodology as described previously [Zhong S. et al. High Throughput Illumina Strand Specific RNA Sequencing Library Preparation. Cold Spring Harb. Protoc. 2011 940 949 2011],wt rep1,SAMEA7579965,Garvan Institute of Medical Research,ENA first public:2020 12 01|ENA last update:2020 11 17|External Id:SAMEA7579965|INSDC center alias:Garvan Institute of Medical Research|INSDC center name:Garvan Institute of Medical Research|INSDC first public:2020 12 01T04:12:08Z|INSDC last update:2020 11 17T14:25:39Z|INSDC status:public|Submitter Id:E MTAB 9777:wt rep1|age:24hpf|broker name:ArrayExpress|common name:zebrafish|developmental stage:embryo stage|organism part:whole organism|sample name:E MTAB 9777:wt rep1|strain:AB / Tubingen,,,,,,,,,Illumina HiSeq 1500 sequencing; Depletion of Foxk transcription factors causes genome wide transcriptional misregulation and developmental arrest in zebrafish embryos,E MTAB 9777:wt rep1 s,wt rep1 s,Depletion of Foxk transcription factors causes genome wide transcriptional misregulation and developmental arrest in zebrafish embryos,Embryos were collected 0 hpf and incubated in 1X E3 medium 0.03% NaCl 0.005% CaCl2 0.0013% KCl 99.9557% H2O 0.008% H14MgO11S for 24 hours at 28.5°C Adult wild type AB/Tubingen Danio rerio zebrafish were bred in a 1 male:1 female ratio. 1c embryos were injected with morpholino oligonucleotides targeting foxk1 foxk2 and foxk2 1 transcripts Total RNA was purified from xxx embryos using the RNeasy Mini Kit Qiagen Valencia CA USA according to the manufacturer's instructions. mRNA Seq libraries were generated from total RNA with polyA+ selection of mRNA using the TruSeq RNA Sample Prep Kit v2 Illumina San Diego CA. Strand specific libraries were constructed using a dUTP methodology as described previously [Zhong S. et al. High Throughput Illumina Strand Specific RNA Sequencing Library Preparation. Cold Spring Harb. Protoc. 2011 940 949 2011],Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 1500,,ERP125162,Illumina HiSeq 1500 sequencing; Depletion of Foxk transcription factors causes genome wide transcriptional misregulation and developmental arrest in zebrafish embryos,ENA FIRST PUBLIC:2020 12 01|ENA LAST UPDATE:2020 11 17,wt_rep1.fastq.gz,fastq,5640822932.0,55849732.0,E MTAB 9777:wt rep1,0:101 1:0,A:1394470185;C:1346058952;G:1354472388;T:1538701573;N:7119834,101,0,,,1394470185,1346058952,1354472388,1538701573,7119834,ERX4714624,ERS5338301,ERA3145932,European Nucleotide Archive,European Nucleotide Archive,1,0.94371,,0.05678,,0.69664,,0.47233,,101,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Unknown,2020-11-17,Pharyngula,Embryo,Whole Organism,All anatomical structures 10070,ERR4844841,ERX4714623,ERS5338300,ERP125162,PRJEB41393,Depletion of Foxk transcription factors causes genome wide transcriptional misregulation and developmental arrest in zebrafish embryos,E-MTAB-9777,Transcriptome Analysis,Foxk proteins are transcriptional regulators implicated in key biological processes such as glycolysis autophagy and cell cycle regulation among others. Here we employ targeted morpholino knockdown to deplete Foxk1 Fokx2 and Foxk2 1 proteins in developing zebrafish embryos. We demonstrate that the loss of Foxk transcription factors causes genome wide transcriptional misregulation characterised by upregulation of autophagy related genes and downregulation of cell cycle regulators. The phenotype is embryonic lethal with the majority of embryos not surviving past 24hpf.,ENA FIRST PUBLIC:2020 12 01|ENA LAST UPDATE:2020 11 17,,Protocols: Embryos were collected 0 hpf and incubated in 1X E3 medium 0.03% NaCl 0.005% CaCl2 0.0013% KCl 99.9557% H2O 0.008% H14MgO11S for 24 hours at 28.5°C Adult wild type AB/Tubingen Danio rerio zebrafish were bred in a 1 male:1 female ratio. 1c embryos were injected with morpholino oligonucleotides targeting foxk1 foxk2 and foxk2 1 transcripts Total RNA was purified from xxx embryos using the RNeasy Mini Kit Qiagen Valencia CA USA according to the manufacturer's instructions. mRNA Seq libraries were generated from total RNA with polyA+ selection of mRNA using the TruSeq RNA Sample Prep Kit v2 Illumina San Diego CA. Strand specific libraries were constructed using a dUTP methodology as described previously [Zhong S. et al. High Throughput Illumina Strand Specific RNA Sequencing Library Preparation. Cold Spring Harb. Protoc. 2011 940 949 2011],foxk MO rep2,SAMEA7579964,Garvan Institute of Medical Research,ENA first public:2020 12 01|ENA last update:2020 11 17|External Id:SAMEA7579964|INSDC center alias:Garvan Institute of Medical Research|INSDC center name:Garvan Institute of Medical Research|INSDC first public:2020 12 01T04:12:08Z|INSDC last update:2020 11 17T14:25:39Z|INSDC status:public|Submitter Id:E MTAB 9777:foxk MO rep2|age:24hpf|broker name:ArrayExpress|common name:zebrafish|developmental stage:embryo stage|organism part:whole organism|sample name:E MTAB 9777:foxk MO rep2|strain:AB / Tubingen,,,,,,,,,Illumina HiSeq 1500 sequencing; Depletion of Foxk transcription factors causes genome wide transcriptional misregulation and developmental arrest in zebrafish embryos,E MTAB 9777:foxk MO rep2 s,foxk MO rep2 s,Depletion of Foxk transcription factors causes genome wide transcriptional misregulation and developmental arrest in zebrafish embryos,Embryos were collected 0 hpf and incubated in 1X E3 medium 0.03% NaCl 0.005% CaCl2 0.0013% KCl 99.9557% H2O 0.008% H14MgO11S for 24 hours at 28.5°C Adult wild type AB/Tubingen Danio rerio zebrafish were bred in a 1 male:1 female ratio. 1c embryos were injected with morpholino oligonucleotides targeting foxk1 foxk2 and foxk2 1 transcripts Total RNA was purified from xxx embryos using the RNeasy Mini Kit Qiagen Valencia CA USA according to the manufacturer's instructions. mRNA Seq libraries were generated from total RNA with polyA+ selection of mRNA using the TruSeq RNA Sample Prep Kit v2 Illumina San Diego CA. Strand specific libraries were constructed using a dUTP methodology as described previously [Zhong S. et al. High Throughput Illumina Strand Specific RNA Sequencing Library Preparation. Cold Spring Harb. Protoc. 2011 940 949 2011],Experimental Factor: compound:morpholino against foxk1/foxk2/foxk2 1|Experimental Factor: dose:9,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 1500,,ERP125162,Illumina HiSeq 1500 sequencing; Depletion of Foxk transcription factors causes genome wide transcriptional misregulation and developmental arrest in zebrafish embryos,ENA FIRST PUBLIC:2020 12 01|ENA LAST UPDATE:2020 11 17,foxk1-2_rep2.fastq.gz,fastq,3550070715.0,35149215.0,E MTAB 9777:foxk MO rep2,0:101 1:0,A:901160433;C:835963324;G:840839574;T:967604789;N:4502595,101,0,,,901160433,835963324,840839574,967604789,4502595,ERX4714623,ERS5338300,ERA3145932,European Nucleotide Archive,European Nucleotide Archive,1,0.94006,,0.07603,,0.69455,,0.46867,,101,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Unknown,2020-11-17,Pharyngula,Embryo,Whole Organism,All anatomical structures 10071,ERR4844840,ERX4714622,ERS5338299,ERP125162,PRJEB41393,Depletion of Foxk transcription factors causes genome wide transcriptional misregulation and developmental arrest in zebrafish embryos,E-MTAB-9777,Transcriptome Analysis,Foxk proteins are transcriptional regulators implicated in key biological processes such as glycolysis autophagy and cell cycle regulation among others. Here we employ targeted morpholino knockdown to deplete Foxk1 Fokx2 and Foxk2 1 proteins in developing zebrafish embryos. We demonstrate that the loss of Foxk transcription factors causes genome wide transcriptional misregulation characterised by upregulation of autophagy related genes and downregulation of cell cycle regulators. The phenotype is embryonic lethal with the majority of embryos not surviving past 24hpf.,ENA FIRST PUBLIC:2020 12 01|ENA LAST UPDATE:2020 11 17,,Protocols: Embryos were collected 0 hpf and incubated in 1X E3 medium 0.03% NaCl 0.005% CaCl2 0.0013% KCl 99.9557% H2O 0.008% H14MgO11S for 24 hours at 28.5°C Adult wild type AB/Tubingen Danio rerio zebrafish were bred in a 1 male:1 female ratio. 1c embryos were injected with morpholino oligonucleotides targeting foxk1 foxk2 and foxk2 1 transcripts Total RNA was purified from xxx embryos using the RNeasy Mini Kit Qiagen Valencia CA USA according to the manufacturer's instructions. mRNA Seq libraries were generated from total RNA with polyA+ selection of mRNA using the TruSeq RNA Sample Prep Kit v2 Illumina San Diego CA. Strand specific libraries were constructed using a dUTP methodology as described previously [Zhong S. et al. High Throughput Illumina Strand Specific RNA Sequencing Library Preparation. Cold Spring Harb. Protoc. 2011 940 949 2011],foxk MO rep1,SAMEA7579963,Garvan Institute of Medical Research,ENA first public:2020 12 01|ENA last update:2020 11 17|External Id:SAMEA7579963|INSDC center alias:Garvan Institute of Medical Research|INSDC center name:Garvan Institute of Medical Research|INSDC first public:2020 12 01T04:12:08Z|INSDC last update:2020 11 17T14:25:39Z|INSDC status:public|Submitter Id:E MTAB 9777:foxk MO rep1|age:24hpf|broker name:ArrayExpress|common name:zebrafish|developmental stage:embryo stage|organism part:whole organism|sample name:E MTAB 9777:foxk MO rep1|strain:AB / Tubingen,,,,,,,,,Illumina HiSeq 1500 sequencing; Depletion of Foxk transcription factors causes genome wide transcriptional misregulation and developmental arrest in zebrafish embryos,E MTAB 9777:foxk MO rep1 s,foxk MO rep1 s,Depletion of Foxk transcription factors causes genome wide transcriptional misregulation and developmental arrest in zebrafish embryos,Embryos were collected 0 hpf and incubated in 1X E3 medium 0.03% NaCl 0.005% CaCl2 0.0013% KCl 99.9557% H2O 0.008% H14MgO11S for 24 hours at 28.5°C Adult wild type AB/Tubingen Danio rerio zebrafish were bred in a 1 male:1 female ratio. 1c embryos were injected with morpholino oligonucleotides targeting foxk1 foxk2 and foxk2 1 transcripts Total RNA was purified from xxx embryos using the RNeasy Mini Kit Qiagen Valencia CA USA according to the manufacturer's instructions. mRNA Seq libraries were generated from total RNA with polyA+ selection of mRNA using the TruSeq RNA Sample Prep Kit v2 Illumina San Diego CA. Strand specific libraries were constructed using a dUTP methodology as described previously [Zhong S. et al. High Throughput Illumina Strand Specific RNA Sequencing Library Preparation. Cold Spring Harb. Protoc. 2011 940 949 2011],Experimental Factor: compound:morpholino against foxk1/foxk2/foxk2 1|Experimental Factor: dose:9,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 1500,,ERP125162,Illumina HiSeq 1500 sequencing; Depletion of Foxk transcription factors causes genome wide transcriptional misregulation and developmental arrest in zebrafish embryos,ENA FIRST PUBLIC:2020 12 01|ENA LAST UPDATE:2020 11 17,foxk1-2_rep1.fastq.gz,fastq,3601987139.0,35663239.0,E MTAB 9777:foxk MO rep1,0:101 1:0,A:910641017;C:846039018;G:853351642;T:987409698;N:4545764,101,0,,,910641017,846039018,853351642,987409698,4545764,ERX4714622,ERS5338299,ERA3145932,European Nucleotide Archive,European Nucleotide Archive,1,0.93784,,0.08061,,0.69292,,0.47364,,101,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Unknown,2020-11-17,Pharyngula,Embryo,Whole Organism,All anatomical structures 10072,ERR4910331,ERX4777154,ERS5435101,ERP125509,PRJEB41694,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,E-MTAB-9853,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used organophosphate insecticide Chlorpyrifos CAS 2921 88 2. The Insecticide Resistance Action Committee IRAC classified Chlorpyrifos post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1B. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of 6PTU for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.75 mg/L high exposure HE 3 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: Catalogue no. 2921 88 2. For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Chlorpyrifos CAS 2921 88 2 PESTANAL® analytical standard was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R842,SAMEA7678119,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678119|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9853:R842|age:96|batch:T10|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:09.12.2019|genotype:wild type genotype|organism part:whole organism|rin:10|sample name:E MTAB 9853:R842|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,E MTAB 9853:R842 s,R842 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,Catalogue no. 2921 88 2. For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Chlorpyrifos CAS 2921 88 2 PESTANAL® analytical standard was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:chlorpyrifos|Experimental Factor: dose:3|Experimental Factor: growth condition:high exposure,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125509,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,R842.fastq.gz,fastq,2078761529.0,41168286.0,E MTAB 9853:R842,0:50.49 1:0,A:525043812;C:513138389;G:493165128;T:542354255;N:5059945,50,0,,,525043812,513138389,493165128,542354255,5059945,ERX4777154,ERS5435101,ERA3184347,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93137,,0.09995,,0.66634,,0.48015,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10073,ERR4910330,ERX4777153,ERS5435100,ERP125509,PRJEB41694,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,E-MTAB-9853,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used organophosphate insecticide Chlorpyrifos CAS 2921 88 2. The Insecticide Resistance Action Committee IRAC classified Chlorpyrifos post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1B. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of 6PTU for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.75 mg/L high exposure HE 3 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: Catalogue no. 2921 88 2. For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Chlorpyrifos CAS 2921 88 2 PESTANAL® analytical standard was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R840,SAMEA7678118,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678118|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9853:R840|age:96|batch:T10|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:09.12.2019|genotype:wild type genotype|organism part:whole organism|rin:10|sample name:E MTAB 9853:R840|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,E MTAB 9853:R840 s,R840 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,Catalogue no. 2921 88 2. For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Chlorpyrifos CAS 2921 88 2 PESTANAL® analytical standard was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:chlorpyrifos|Experimental Factor: dose:0.00075|Experimental Factor: growth condition:low exposure,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125509,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,R840.fastq.gz,fastq,2344231087.0,46394848.0,E MTAB 9853:R840,0:50.53 1:0,A:603829684;C:570903008;G:545594153;T:621405057;N:2499185,50,0,,,603829684,570903008,545594153,621405057,2499185,ERX4777153,ERS5435100,ERA3184347,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.92792,,0.10519,,0.6646,,0.48351,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10074,ERR4910329,ERX4777152,ERS5435099,ERP125509,PRJEB41694,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,E-MTAB-9853,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used organophosphate insecticide Chlorpyrifos CAS 2921 88 2. The Insecticide Resistance Action Committee IRAC classified Chlorpyrifos post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1B. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of 6PTU for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.75 mg/L high exposure HE 3 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: Catalogue no. 2921 88 2. For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Chlorpyrifos CAS 2921 88 2 PESTANAL® analytical standard was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R839,SAMEA7678117,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678117|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9853:R839|age:96|batch:T10|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:09.12.2019|genotype:wild type genotype|organism part:whole organism|rin:10|sample name:E MTAB 9853:R839|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,E MTAB 9853:R839 s,R839 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,Catalogue no. 2921 88 2. For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Chlorpyrifos CAS 2921 88 2 PESTANAL® analytical standard was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:n1|Experimental Factor: growth condition:normal,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125509,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,R839.fastq.gz,fastq,1996290916.0,39508020.0,E MTAB 9853:R839,0:50.53 1:0,A:509362981;C:490497777;G:469414712;T:524505312;N:2510134,50,0,,,509362981,490497777,469414712,524505312,2510134,ERX4777152,ERS5435099,ERA3184347,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93138,,0.10299,,0.66468,,0.48637,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10075,ERR4910328,ERX4777151,ERS5435098,ERP125509,PRJEB41694,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,E-MTAB-9853,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used organophosphate insecticide Chlorpyrifos CAS 2921 88 2. The Insecticide Resistance Action Committee IRAC classified Chlorpyrifos post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1B. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of 6PTU for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.75 mg/L high exposure HE 3 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: Catalogue no. 2921 88 2. For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Chlorpyrifos CAS 2921 88 2 PESTANAL® analytical standard was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R838,SAMEA7678116,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678116|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9853:R838|age:96|batch:T2|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:09.12.2019|genotype:wild type genotype|organism part:whole organism|rin:9 9|sample name:E MTAB 9853:R838|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,E MTAB 9853:R838 s,R838 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,Catalogue no. 2921 88 2. For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Chlorpyrifos CAS 2921 88 2 PESTANAL® analytical standard was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:chlorpyrifos|Experimental Factor: dose:3|Experimental Factor: growth condition:high exposure,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125509,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,R838.fastq.gz,fastq,1801740137.0,35669320.0,E MTAB 9853:R838,0:50.51 1:0,A:466977493;C:436158087;G:416333880;T:479487201;N:2783476,50,0,,,466977493,436158087,416333880,479487201,2783476,ERX4777151,ERS5435098,ERA3184347,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.92647,,0.11533,,0.666,,0.48331,,49,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10076,ERR4910327,ERX4777150,ERS5435097,ERP125509,PRJEB41694,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,E-MTAB-9853,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used organophosphate insecticide Chlorpyrifos CAS 2921 88 2. The Insecticide Resistance Action Committee IRAC classified Chlorpyrifos post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1B. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of 6PTU for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.75 mg/L high exposure HE 3 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: Catalogue no. 2921 88 2. For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Chlorpyrifos CAS 2921 88 2 PESTANAL® analytical standard was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R836,SAMEA7678115,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678115|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9853:R836|age:96|batch:T2|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:09.12.2019|genotype:wild type genotype|organism part:whole organism|rin:9 8|sample name:E MTAB 9853:R836|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,E MTAB 9853:R836 s,R836 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,Catalogue no. 2921 88 2. For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Chlorpyrifos CAS 2921 88 2 PESTANAL® analytical standard was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:chlorpyrifos|Experimental Factor: dose:0.00075|Experimental Factor: growth condition:low exposure,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125509,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,R836.fastq.gz,fastq,1816994468.0,35959155.0,E MTAB 9853:R836,0:50.53 1:0,A:465931946;C:444466278;G:425419784;T:479276121;N:1900339,50,0,,,465931946,444466278,425419784,479276121,1900339,ERX4777150,ERS5435097,ERA3184347,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.92973,,0.10412,,0.66425,,0.4798,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10077,ERR4910326,ERX4777149,ERS5435096,ERP125509,PRJEB41694,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,E-MTAB-9853,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used organophosphate insecticide Chlorpyrifos CAS 2921 88 2. The Insecticide Resistance Action Committee IRAC classified Chlorpyrifos post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1B. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of 6PTU for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.75 mg/L high exposure HE 3 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: Catalogue no. 2921 88 2. For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Chlorpyrifos CAS 2921 88 2 PESTANAL® analytical standard was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R835,SAMEA7678114,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678114|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9853:R835|age:96|batch:T2|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:09.12.2019|genotype:wild type genotype|organism part:whole organism|rin:9 9|sample name:E MTAB 9853:R835|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,E MTAB 9853:R835 s,R835 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,Catalogue no. 2921 88 2. For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Chlorpyrifos CAS 2921 88 2 PESTANAL® analytical standard was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:n1|Experimental Factor: growth condition:normal,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125509,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,R835.fastq.gz,fastq,2309123342.0,45698247.0,E MTAB 9853:R835,0:50.53 1:0,A:590448588;C:564838103;G:542833223;T:608521072;N:2482356,50,0,,,590448588,564838103,542833223,608521072,2482356,ERX4777149,ERS5435096,ERA3184347,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.92963,,0.10535,,0.66156,,0.48118,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10078,ERR4910325,ERX4777148,ERS5435095,ERP125509,PRJEB41694,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,E-MTAB-9853,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used organophosphate insecticide Chlorpyrifos CAS 2921 88 2. The Insecticide Resistance Action Committee IRAC classified Chlorpyrifos post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1B. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of 6PTU for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.75 mg/L high exposure HE 3 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: Catalogue no. 2921 88 2. For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Chlorpyrifos CAS 2921 88 2 PESTANAL® analytical standard was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R834,SAMEA7678113,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678113|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9853:R834|age:96|batch:T11|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:09.12.2019|genotype:wild type genotype|organism part:whole organism|rin:9 8|sample name:E MTAB 9853:R834|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,E MTAB 9853:R834 s,R834 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,Catalogue no. 2921 88 2. For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Chlorpyrifos CAS 2921 88 2 PESTANAL® analytical standard was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:chlorpyrifos|Experimental Factor: dose:3|Experimental Factor: growth condition:high exposure,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125509,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,R834.fastq.gz,fastq,2122701060.0,42013381.0,E MTAB 9853:R834,0:50.52 1:0,A:547558397;C:515792741;G:494125094;T:562812656;N:2412172,50,0,,,547558397,515792741,494125094,562812656,2412172,ERX4777148,ERS5435095,ERA3184347,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.92823,,0.11135,,0.66492,,0.48309,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10079,ERR4910324,ERX4777147,ERS5435094,ERP125509,PRJEB41694,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,E-MTAB-9853,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used organophosphate insecticide Chlorpyrifos CAS 2921 88 2. The Insecticide Resistance Action Committee IRAC classified Chlorpyrifos post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1B. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of 6PTU for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.75 mg/L high exposure HE 3 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: Catalogue no. 2921 88 2. For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Chlorpyrifos CAS 2921 88 2 PESTANAL® analytical standard was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R832,SAMEA7678112,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678112|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9853:R832|age:96|batch:T11|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:09.12.2019|genotype:wild type genotype|organism part:whole organism|rin:9 7|sample name:E MTAB 9853:R832|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,E MTAB 9853:R832 s,R832 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,Catalogue no. 2921 88 2. For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Chlorpyrifos CAS 2921 88 2 PESTANAL® analytical standard was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:chlorpyrifos|Experimental Factor: dose:0.00075|Experimental Factor: growth condition:low exposure,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125509,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,R832.fastq.gz,fastq,2407016662.0,47713826.0,E MTAB 9853:R832,0:50.45 1:0,A:637497754;C:575428535;G:531202727;T:659395837;N:3491809,50,0,,,637497754,575428535,531202727,659395837,3491809,ERX4777147,ERS5435094,ERA3184347,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.91708,,0.12568,,0.65494,,0.48103,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10080,ERR4910323,ERX4777146,ERS5435093,ERP125509,PRJEB41694,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,E-MTAB-9853,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used organophosphate insecticide Chlorpyrifos CAS 2921 88 2. The Insecticide Resistance Action Committee IRAC classified Chlorpyrifos post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1B. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of 6PTU for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.75 mg/L high exposure HE 3 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: Catalogue no. 2921 88 2. For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Chlorpyrifos CAS 2921 88 2 PESTANAL® analytical standard was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R831,SAMEA7678111,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678111|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9853:R831|age:96|batch:T11|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:09.12.2019|genotype:wild type genotype|organism part:whole organism|rin:9 9|sample name:E MTAB 9853:R831|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,E MTAB 9853:R831 s,R831 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,Catalogue no. 2921 88 2. For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Chlorpyrifos CAS 2921 88 2 PESTANAL® analytical standard was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:n1|Experimental Factor: growth condition:normal,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125509,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Chlorpyrifos below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,R831.fastq.gz,fastq,2084924019.0,41265710.0,E MTAB 9853:R831,0:50.52 1:0,A:529532094;C:513298404;G:493028485;T:546325575;N:2739461,50,0,,,529532094,513298404,493028485,546325575,2739461,ERX4777146,ERS5435093,ERA3184347,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93121,,0.10702,,0.66064,,0.47779,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10081,ERR4910712,ERX4777535,ERS5435303,ERP125510,PRJEB41695,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E-MTAB-9852,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of Abamectin CAS 71751 41 2. Abamectin is a heavily used insecticide applied for crop protection against sucking insects i.e. Acari. The Insecticide Resistance Action Committee IRAC classified Abamectin post its mode of action MoA in the target organism as a Glutamate gated chloride channel GluCl allosteric modulator Group 6. In vertebrates GluCl do not exist but they are closely related to vertebrate glycine receptors Wolstenholme 2012. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Abamectin for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.11 mg/L mid exposure ME 0.22 mg/L high exposure HE 0.44 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R249,SAMEA7678322,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678322|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9852:R249|age:96|batch:T14 3|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:18.02.2019|genotype:wild type genotype|individual:mixed pool of 10 larvae|organism part:whole organism|rin:9 7|sample name:E MTAB 9852:R249|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E MTAB 9852:R249 s,R249 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: growth condition:mid exposure|Experimental Factor: compound:abamectin|Experimental Factor: dose:0.00022,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125510,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p823sR249_sr.fastq.gz,fastq,1238386140.0,24550785.0,E MTAB 9852:R249,0:50.44 1:0,A:315619803;C:300278943;G:290065564;T:328018594;N:4403236,50,0,,,315619803,300278943,290065564,328018594,4403236,ERX4777535,ERS5435303,ERA3184562,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.94195,,0.10394,,0.65249,,0.46521,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10082,ERR4910711,ERX4777534,ERS5435302,ERP125510,PRJEB41695,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E-MTAB-9852,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of Abamectin CAS 71751 41 2. Abamectin is a heavily used insecticide applied for crop protection against sucking insects i.e. Acari. The Insecticide Resistance Action Committee IRAC classified Abamectin post its mode of action MoA in the target organism as a Glutamate gated chloride channel GluCl allosteric modulator Group 6. In vertebrates GluCl do not exist but they are closely related to vertebrate glycine receptors Wolstenholme 2012. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Abamectin for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.11 mg/L mid exposure ME 0.22 mg/L high exposure HE 0.44 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R243,SAMEA7678321,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678321|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9852:R243|age:96|batch:T8|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:11.02.2019|genotype:wild type genotype|individual:mixed pool of 10 larvae|organism part:whole organism|rin:9 8|sample name:E MTAB 9852:R243|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E MTAB 9852:R243 s,R243 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: growth condition:mid exposure|Experimental Factor: compound:abamectin|Experimental Factor: dose:0.00022,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125510,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p823sR243_sr.fastq.gz,fastq,2421445373.0,47945201.0,E MTAB 9852:R243,0:50.50 1:0,A:644050981;C:566576386;G:556332105;T:652514946;N:1970955,50,0,,,644050981,566576386,556332105,652514946,1970955,ERX4777534,ERS5435302,ERA3184562,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93336,,0.11777,,0.64881,,0.47801,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10083,ERR4910710,ERX4777533,ERS5435301,ERP125510,PRJEB41695,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E-MTAB-9852,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of Abamectin CAS 71751 41 2. Abamectin is a heavily used insecticide applied for crop protection against sucking insects i.e. Acari. The Insecticide Resistance Action Committee IRAC classified Abamectin post its mode of action MoA in the target organism as a Glutamate gated chloride channel GluCl allosteric modulator Group 6. In vertebrates GluCl do not exist but they are closely related to vertebrate glycine receptors Wolstenholme 2012. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Abamectin for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.11 mg/L mid exposure ME 0.22 mg/L high exposure HE 0.44 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R237,SAMEA7678320,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678320|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9852:R237|age:96|batch:T6|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:11.02.2019|genotype:wild type genotype|individual:mixed pool of 10 larvae|organism part:whole organism|rin:10|sample name:E MTAB 9852:R237|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E MTAB 9852:R237 s,R237 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: growth condition:mid exposure|Experimental Factor: compound:abamectin|Experimental Factor: dose:0.00022,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125510,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p823sR237_sr.fastq.gz,fastq,1226270967.0,24309553.0,E MTAB 9852:R237,0:50.44 1:0,A:315571166;C:295129287;G:284455226;T:326832027;N:4283261,50,0,,,315571166,295129287,284455226,326832027,4283261,ERX4777533,ERS5435301,ERA3184562,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.941,,0.11242,,0.65504,,0.47821,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10084,ERR4910709,ERX4777532,ERS5435300,ERP125510,PRJEB41695,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E-MTAB-9852,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of Abamectin CAS 71751 41 2. Abamectin is a heavily used insecticide applied for crop protection against sucking insects i.e. Acari. The Insecticide Resistance Action Committee IRAC classified Abamectin post its mode of action MoA in the target organism as a Glutamate gated chloride channel GluCl allosteric modulator Group 6. In vertebrates GluCl do not exist but they are closely related to vertebrate glycine receptors Wolstenholme 2012. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Abamectin for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.11 mg/L mid exposure ME 0.22 mg/L high exposure HE 0.44 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R248,SAMEA7678319,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678319|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9852:R248|age:96|batch:T14 3|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:18.02.2019|genotype:wild type genotype|individual:mixed pool of 10 larvae|organism part:whole organism|rin:9 4|sample name:E MTAB 9852:R248|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E MTAB 9852:R248 s,R248 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: growth condition:low exposure|Experimental Factor: compound:abamectin|Experimental Factor: dose:0.00011,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125510,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p689sR248_sr.fastq.gz,fastq,1418368154.0,28100223.0,E MTAB 9852:R248,0:50.48 1:0,A:365721817;C:343904004;G:327397507;T:379298412;N:2046414,50,0,,,365721817,343904004,327397507,379298412,2046414,ERX4777532,ERS5435300,ERA3184562,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93616,,0.10439,,0.65502,,0.47312,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10085,ERR4910708,ERX4777531,ERS5435299,ERP125510,PRJEB41695,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E-MTAB-9852,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of Abamectin CAS 71751 41 2. Abamectin is a heavily used insecticide applied for crop protection against sucking insects i.e. Acari. The Insecticide Resistance Action Committee IRAC classified Abamectin post its mode of action MoA in the target organism as a Glutamate gated chloride channel GluCl allosteric modulator Group 6. In vertebrates GluCl do not exist but they are closely related to vertebrate glycine receptors Wolstenholme 2012. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Abamectin for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.11 mg/L mid exposure ME 0.22 mg/L high exposure HE 0.44 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R242,SAMEA7678318,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678318|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9852:R242|age:96|batch:T8|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:11.02.2019|genotype:wild type genotype|individual:mixed pool of 10 larvae|organism part:whole organism|rin:10|sample name:E MTAB 9852:R242|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E MTAB 9852:R242 s,R242 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: growth condition:low exposure|Experimental Factor: compound:abamectin|Experimental Factor: dose:0.00011,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125510,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p689sR242_sr.fastq.gz,fastq,1078378173.0,21357646.0,E MTAB 9852:R242,0:50.49 1:0,A:279746568;C:260936637;G:248227414;T:288521601;N:945953,50,0,,,279746568,260936637,248227414,288521601,945953,ERX4777531,ERS5435299,ERA3184562,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.9353,,0.11226,,0.6552,,0.47905,,49,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10086,ERR4910707,ERX4777530,ERS5435298,ERP125510,PRJEB41695,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E-MTAB-9852,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of Abamectin CAS 71751 41 2. Abamectin is a heavily used insecticide applied for crop protection against sucking insects i.e. Acari. The Insecticide Resistance Action Committee IRAC classified Abamectin post its mode of action MoA in the target organism as a Glutamate gated chloride channel GluCl allosteric modulator Group 6. In vertebrates GluCl do not exist but they are closely related to vertebrate glycine receptors Wolstenholme 2012. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Abamectin for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.11 mg/L mid exposure ME 0.22 mg/L high exposure HE 0.44 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R236,SAMEA7678317,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678317|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9852:R236|age:96|batch:T6|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:11.02.2019|genotype:wild type genotype|individual:mixed pool of 10 larvae|organism part:whole organism|rin:10|sample name:E MTAB 9852:R236|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E MTAB 9852:R236 s,R236 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: growth condition:low exposure|Experimental Factor: compound:abamectin|Experimental Factor: dose:0.00011,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125510,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p689sR236_sr.fastq.gz,fastq,1095647196.0,21699085.0,E MTAB 9852:R236,0:50.49 1:0,A:284611814;C:265266246;G:251286658;T:293543067;N:939411,50,0,,,284611814,265266246,251286658,293543067,939411,ERX4777530,ERS5435298,ERA3184562,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93368,,0.11269,,0.65748,,0.47474,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10087,ERR4910706,ERX4777529,ERS5435297,ERP125510,PRJEB41695,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E-MTAB-9852,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of Abamectin CAS 71751 41 2. Abamectin is a heavily used insecticide applied for crop protection against sucking insects i.e. Acari. The Insecticide Resistance Action Committee IRAC classified Abamectin post its mode of action MoA in the target organism as a Glutamate gated chloride channel GluCl allosteric modulator Group 6. In vertebrates GluCl do not exist but they are closely related to vertebrate glycine receptors Wolstenholme 2012. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Abamectin for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.11 mg/L mid exposure ME 0.22 mg/L high exposure HE 0.44 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R250,SAMEA7678316,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678316|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9852:R250|age:96|batch:T14 3|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:18.02.2019|genotype:wild type genotype|individual:mixed pool of 10 larvae|organism part:whole organism|rin:9 3|sample name:E MTAB 9852:R250|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E MTAB 9852:R250 s,R250 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: growth condition:high exposure|Experimental Factor: compound:abamectin|Experimental Factor: dose:0.00044,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125510,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p689sR250_sr.fastq.gz,fastq,1360394339.0,26941236.0,E MTAB 9852:R250,0:50.49 1:0,A:350782395;C:329394637;G:315556205;T:363589748;N:1071354,50,0,,,350782395,329394637,315556205,363589748,1071354,ERX4777529,ERS5435297,ERA3184562,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93459,,0.1084,,0.65612,,0.46181,,49,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10088,ERR4910705,ERX4777528,ERS5435296,ERP125510,PRJEB41695,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E-MTAB-9852,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of Abamectin CAS 71751 41 2. Abamectin is a heavily used insecticide applied for crop protection against sucking insects i.e. Acari. The Insecticide Resistance Action Committee IRAC classified Abamectin post its mode of action MoA in the target organism as a Glutamate gated chloride channel GluCl allosteric modulator Group 6. In vertebrates GluCl do not exist but they are closely related to vertebrate glycine receptors Wolstenholme 2012. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Abamectin for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.11 mg/L mid exposure ME 0.22 mg/L high exposure HE 0.44 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R245,SAMEA7678315,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678315|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9852:R245|age:96|batch:T8|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:11.02.2019|genotype:wild type genotype|individual:mixed pool of 10 larvae|organism part:whole organism|rin:9 6|sample name:E MTAB 9852:R245|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E MTAB 9852:R245 s,R245 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: growth condition:high exposure|Experimental Factor: compound:abamectin|Experimental Factor: dose:0.00044,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125510,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p689sR245_sr.fastq.gz,fastq,4133671592.0,81841196.0,E MTAB 9852:R245,0:50.51 1:0,A:1069364626;C:992843654;G:956238531;T:1113501572;N:1723209,50,0,,,1069364626,992843654,956238531,1113501572,1723209,ERX4777528,ERS5435296,ERA3184562,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93652,,0.11134,,0.65196,,0.4699,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10089,ERR4910704,ERX4777527,ERS5435295,ERP125510,PRJEB41695,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E-MTAB-9852,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of Abamectin CAS 71751 41 2. Abamectin is a heavily used insecticide applied for crop protection against sucking insects i.e. Acari. The Insecticide Resistance Action Committee IRAC classified Abamectin post its mode of action MoA in the target organism as a Glutamate gated chloride channel GluCl allosteric modulator Group 6. In vertebrates GluCl do not exist but they are closely related to vertebrate glycine receptors Wolstenholme 2012. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Abamectin for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.11 mg/L mid exposure ME 0.22 mg/L high exposure HE 0.44 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R239,SAMEA7678314,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678314|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9852:R239|age:96|batch:T6|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:11.02.2019|genotype:wild type genotype|individual:mixed pool of 10 larvae|organism part:whole organism|rin:9 8|sample name:E MTAB 9852:R239|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E MTAB 9852:R239 s,R239 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: growth condition:high exposure|Experimental Factor: compound:abamectin|Experimental Factor: dose:0.00044,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125510,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p689sR239_sr.fastq.gz,fastq,1265182243.0,25065223.0,E MTAB 9852:R239,0:50.48 1:0,A:328276812;C:305137622;G:290542708;T:339548031;N:1677070,50,0,,,328276812,305137622,290542708,339548031,1677070,ERX4777527,ERS5435295,ERA3184562,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93333,,0.11548,,0.65354,,0.47419,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10090,ERR4910703,ERX4777526,ERS5435294,ERP125510,PRJEB41695,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E-MTAB-9852,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of Abamectin CAS 71751 41 2. Abamectin is a heavily used insecticide applied for crop protection against sucking insects i.e. Acari. The Insecticide Resistance Action Committee IRAC classified Abamectin post its mode of action MoA in the target organism as a Glutamate gated chloride channel GluCl allosteric modulator Group 6. In vertebrates GluCl do not exist but they are closely related to vertebrate glycine receptors Wolstenholme 2012. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Abamectin for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.11 mg/L mid exposure ME 0.22 mg/L high exposure HE 0.44 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R247,SAMEA7678313,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678313|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9852:R247|age:96|batch:T14 3|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:18.02.2019|genotype:wild type genotype|individual:mixed pool of 10 larvae|organism part:whole organism|rin:9 7|sample name:E MTAB 9852:R247|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E MTAB 9852:R247 s,R247 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: growth condition:normal|Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125510,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p689sR247_sr.fastq.gz,fastq,1537094317.0,30449470.0,E MTAB 9852:R247,0:50.48 1:0,A:378609932;C:387657971;G:375197386;T:392878686;N:2750342,50,0,,,378609932,387657971,375197386,392878686,2750342,ERX4777526,ERS5435294,ERA3184562,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.94344,,0.08094,,0.66093,,0.47759,,49,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10091,ERR4910702,ERX4777525,ERS5435293,ERP125510,PRJEB41695,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E-MTAB-9852,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of Abamectin CAS 71751 41 2. Abamectin is a heavily used insecticide applied for crop protection against sucking insects i.e. Acari. The Insecticide Resistance Action Committee IRAC classified Abamectin post its mode of action MoA in the target organism as a Glutamate gated chloride channel GluCl allosteric modulator Group 6. In vertebrates GluCl do not exist but they are closely related to vertebrate glycine receptors Wolstenholme 2012. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Abamectin for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.11 mg/L mid exposure ME 0.22 mg/L high exposure HE 0.44 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R241,SAMEA7678312,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678312|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9852:R241|age:96|batch:T8|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:11.02.2019|genotype:wild type genotype|individual:mixed pool of 10 larvae|organism part:whole organism|rin:9 7|sample name:E MTAB 9852:R241|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E MTAB 9852:R241 s,R241 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: growth condition:normal|Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125510,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p823sR241_sr.fastq.gz,fastq,1421173155.0,28241274.0,E MTAB 9852:R241,0:50.32 1:0,A:352339224;C:349259614;G:337822083;T:368520360;N:13231874,50,0,,,352339224,349259614,337822083,368520360,13231874,ERX4777525,ERS5435293,ERA3184562,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.94609,,0.08553,,0.65729,,0.47079,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10092,ERR4910701,ERX4777524,ERS5435292,ERP125510,PRJEB41695,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E-MTAB-9852,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of Abamectin CAS 71751 41 2. Abamectin is a heavily used insecticide applied for crop protection against sucking insects i.e. Acari. The Insecticide Resistance Action Committee IRAC classified Abamectin post its mode of action MoA in the target organism as a Glutamate gated chloride channel GluCl allosteric modulator Group 6. In vertebrates GluCl do not exist but they are closely related to vertebrate glycine receptors Wolstenholme 2012. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Abamectin for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.11 mg/L mid exposure ME 0.22 mg/L high exposure HE 0.44 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R235,SAMEA7678311,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678311|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9852:R235|age:96|batch:T6|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|extraction date:11.02.2019|genotype:wild type genotype|individual:mixed pool of 10 larvae|organism part:whole organism|rin:10|sample name:E MTAB 9852:R235|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,E MTAB 9852:R235 s,R235 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. CASRN: 71751 41 2 Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Abamectin CAS: 71751 41 2 PESTANAL© analytical grade was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. Then 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: growth condition:normal|Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125510,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Abamectin below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p689sR235_sr.fastq.gz,fastq,1260969924.0,24967845.0,E MTAB 9852:R235,0:50.50 1:0,A:321334927;C:309076905;G:296926662;T:332736920;N:894510,50,0,,,321334927,309076905,296926662,332736920,894510,ERX4777524,ERS5435292,ERA3184562,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.94005,,0.10804,,0.66014,,0.48173,,49,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10093,ERR4910723,ERX4777546,ERS5435312,ERP125511,PRJEB41696,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,E-MTAB-9854,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used organophosphate insecticide Fipronil CAS 2921 88 2. The Insecticide Resistance Action Committee IRAC classified Fipronil post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1B. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Fipronil for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.075 mg/L high exposure HE 0.3 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Fipronil CAS 120068 37 3 TraceCERT® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R852,SAMEA7678331,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678331|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9854:R852|age:96|broker name:ArrayExpress|cas:120068 37 3|common name:zebrafish|condition:low exposure|developmental stage:larval stage|extraction date:27.01.2020|genotype:wild type genotype|nominal conc:0.000075|organism part:whole organism|pooled individuals:10|rin:9.2|sample name:E MTAB 9854:R852|sex:mixed|strain:AB|tank spawning group:T12,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,E MTAB 9854:R852 s,R852 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Fipronil CAS 120068 37 3 TraceCERT® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:fipronil|Experimental Factor: dose:0.075,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125511,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,R852.fastq.gz,fastq,1501461841.0,29758462.0,E MTAB 9854:R852,0:50.45 1:0,A:383178331;C:363921609;G:352164753;T:397820217;N:4376931,50,0,,,383178331,363921609,352164753,397820217,4376931,ERX4777546,ERS5435312,ERA3184565,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.94338,,0.10128,,0.65161,,0.47039,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10094,ERR4910722,ERX4777545,ERS5435311,ERP125511,PRJEB41696,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,E-MTAB-9854,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used organophosphate insecticide Fipronil CAS 2921 88 2. The Insecticide Resistance Action Committee IRAC classified Fipronil post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1B. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Fipronil for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.075 mg/L high exposure HE 0.3 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Fipronil CAS 120068 37 3 TraceCERT® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R848,SAMEA7678330,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678330|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9854:R848|age:96|broker name:ArrayExpress|cas:120068 37 3|common name:zebrafish|condition:low exposure|developmental stage:larval stage|extraction date:27.01.2020|genotype:wild type genotype|nominal conc:0.000075|organism part:whole organism|pooled individuals:10|rin:9.1|sample name:E MTAB 9854:R848|sex:mixed|strain:AB|tank spawning group:T11,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,E MTAB 9854:R848 s,R848 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Fipronil CAS 120068 37 3 TraceCERT® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:fipronil|Experimental Factor: dose:0.075,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125511,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,R848.fastq.gz,fastq,1270434813.0,25300616.0,E MTAB 9854:R848,0:50.21 1:0,A:323517980;C:302337722;G:291425285;T:335522576;N:17631250,50,0,,,323517980,302337722,291425285,335522576,17631250,ERX4777545,ERS5435311,ERA3184565,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93466,,0.10517,,0.65425,,0.48408,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10095,ERR4910721,ERX4777544,ERS5435310,ERP125511,PRJEB41696,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,E-MTAB-9854,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used organophosphate insecticide Fipronil CAS 2921 88 2. The Insecticide Resistance Action Committee IRAC classified Fipronil post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1B. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Fipronil for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.075 mg/L high exposure HE 0.3 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Fipronil CAS 120068 37 3 TraceCERT® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R844,SAMEA7678329,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678329|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9854:R844|age:96|broker name:ArrayExpress|cas:120068 37 3|common name:zebrafish|condition:low exposure|developmental stage:larval stage|extraction date:27.01.2020|genotype:wild type genotype|nominal conc:0.000075|organism part:whole organism|pooled individuals:10|rin:9.7|sample name:E MTAB 9854:R844|sex:mixed|strain:AB|tank spawning group:T10,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,E MTAB 9854:R844 s,R844 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Fipronil CAS 120068 37 3 TraceCERT® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:fipronil|Experimental Factor: dose:0.075,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125511,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,R844.fastq.gz,fastq,1617418458.0,32083973.0,E MTAB 9854:R844,0:50.41 1:0,A:414770839;C:390980511;G:372715594;T:431908857;N:7042657,50,0,,,414770839,390980511,372715594,431908857,7042657,ERX4777544,ERS5435310,ERA3184565,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93517,,0.11298,,0.64999,,0.48035,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10096,ERR4910720,ERX4777543,ERS5435309,ERP125511,PRJEB41696,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,E-MTAB-9854,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used organophosphate insecticide Fipronil CAS 2921 88 2. The Insecticide Resistance Action Committee IRAC classified Fipronil post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1B. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Fipronil for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.075 mg/L high exposure HE 0.3 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Fipronil CAS 120068 37 3 TraceCERT® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R854,SAMEA7678328,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678328|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9854:R854|age:96|broker name:ArrayExpress|cas:120068 37 3|common name:zebrafish|condition:high exposure|developmental stage:larval stage|extraction date:27.01.2020|genotype:wild type genotype|nominal conc:0.0003|organism part:whole organism|pooled individuals:10|rin:9.8|sample name:E MTAB 9854:R854|sex:mixed|strain:AB|tank spawning group:T12,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,E MTAB 9854:R854 s,R854 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Fipronil CAS 120068 37 3 TraceCERT® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:fipronil|Experimental Factor: dose:0.3,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125511,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,R854.fastq.gz,fastq,1673652816.0,33145949.0,E MTAB 9854:R854,0:50.49 1:0,A:424673752;C:411510497;G:395090177;T:440173524;N:2204866,50,0,,,424673752,411510497,395090177,440173524,2204866,ERX4777543,ERS5435309,ERA3184565,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.94329,,0.09755,,0.65642,,0.4654,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10097,ERR4910719,ERX4777542,ERS5435308,ERP125511,PRJEB41696,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,E-MTAB-9854,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used organophosphate insecticide Fipronil CAS 2921 88 2. The Insecticide Resistance Action Committee IRAC classified Fipronil post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1B. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Fipronil for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.075 mg/L high exposure HE 0.3 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Fipronil CAS 120068 37 3 TraceCERT® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R850,SAMEA7678327,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678327|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9854:R850|age:96|broker name:ArrayExpress|cas:120068 37 3|common name:zebrafish|condition:high exposure|developmental stage:larval stage|extraction date:27.01.2020|genotype:wild type genotype|nominal conc:0.0003|organism part:whole organism|pooled individuals:10|rin:9.2|sample name:E MTAB 9854:R850|sex:mixed|strain:AB|tank spawning group:T11,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,E MTAB 9854:R850 s,R850 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Fipronil CAS 120068 37 3 TraceCERT® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:fipronil|Experimental Factor: dose:0.3,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125511,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,R850.fastq.gz,fastq,2896828206.0,57502950.0,E MTAB 9854:R850,0:50.38 1:0,A:733652981;C:704288937;G:676740479;T:762532101;N:19613708,50,0,,,733652981,704288937,676740479,762532101,19613708,ERX4777542,ERS5435308,ERA3184565,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93972,,0.1116,,0.65504,,0.47885,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10098,ERR4910718,ERX4777541,ERS5435307,ERP125511,PRJEB41696,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,E-MTAB-9854,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used organophosphate insecticide Fipronil CAS 2921 88 2. The Insecticide Resistance Action Committee IRAC classified Fipronil post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1B. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Fipronil for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.075 mg/L high exposure HE 0.3 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Fipronil CAS 120068 37 3 TraceCERT® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R846,SAMEA7678326,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678326|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9854:R846|age:96|broker name:ArrayExpress|cas:120068 37 3|common name:zebrafish|condition:high exposure|developmental stage:larval stage|extraction date:27.01.2020|genotype:wild type genotype|nominal conc:0.0003|organism part:whole organism|pooled individuals:10|rin:9|sample name:E MTAB 9854:R846|sex:mixed|strain:AB|tank spawning group:T10,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,E MTAB 9854:R846 s,R846 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Fipronil CAS 120068 37 3 TraceCERT® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:fipronil|Experimental Factor: dose:0.3,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125511,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,R846.fastq.gz,fastq,1287980321.0,25537756.0,E MTAB 9854:R846,0:50.43 1:0,A:327403451;C:313312050;G:302396862;T:339963183;N:4904775,50,0,,,327403451,313312050,302396862,339963183,4904775,ERX4777541,ERS5435307,ERA3184565,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93976,,0.11403,,0.6536,,0.48517,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10099,ERR4910717,ERX4777540,ERS5435306,ERP125511,PRJEB41696,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,E-MTAB-9854,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used organophosphate insecticide Fipronil CAS 2921 88 2. The Insecticide Resistance Action Committee IRAC classified Fipronil post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1B. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Fipronil for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.075 mg/L high exposure HE 0.3 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Fipronil CAS 120068 37 3 TraceCERT® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R851,SAMEA7678325,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678325|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9854:R851|age:96|broker name:ArrayExpress|cas:120068 37 3|common name:zebrafish|condition:control|developmental stage:larval stage|extraction date:27.01.2020|genotype:wild type genotype|nominal conc:0|organism part:whole organism|pooled individuals:10|rin:9|sample name:E MTAB 9854:R851|sex:mixed|strain:AB|tank spawning group:T12,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,E MTAB 9854:R851 s,R851 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Fipronil CAS 120068 37 3 TraceCERT® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125511,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,R851.fastq.gz,fastq,1910586865.0,37884355.0,E MTAB 9854:R851,0:50.43 1:0,A:485650011;C:464135765;G:448149274;T:504995668;N:7656147,50,0,,,485650011,464135765,448149274,504995668,7656147,ERX4777540,ERS5435306,ERA3184565,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93994,,0.0996,,0.65119,,0.47429,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10100,ERR4910716,ERX4777539,ERS5435305,ERP125511,PRJEB41696,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,E-MTAB-9854,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used organophosphate insecticide Fipronil CAS 2921 88 2. The Insecticide Resistance Action Committee IRAC classified Fipronil post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1B. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Fipronil for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.075 mg/L high exposure HE 0.3 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Fipronil CAS 120068 37 3 TraceCERT® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R847,SAMEA7678324,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678324|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9854:R847|age:96|broker name:ArrayExpress|cas:120068 37 3|common name:zebrafish|condition:control|developmental stage:larval stage|extraction date:27.01.2020|genotype:wild type genotype|nominal conc:0|organism part:whole organism|pooled individuals:10|rin:9.1|sample name:E MTAB 9854:R847|sex:mixed|strain:AB|tank spawning group:T11,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,E MTAB 9854:R847 s,R847 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Fipronil CAS 120068 37 3 TraceCERT® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125511,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,R847.fastq.gz,fastq,1843302575.0,36600884.0,E MTAB 9854:R847,0:50.36 1:0,A:468929467;C:444937076;G:429160492;T:486948036;N:13327504,50,0,,,468929467,444937076,429160492,486948036,13327504,ERX4777539,ERS5435305,ERA3184565,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93829,,0.10472,,0.65212,,0.47795,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10101,ERR4910715,ERX4777538,ERS5435304,ERP125511,PRJEB41696,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,E-MTAB-9854,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used organophosphate insecticide Fipronil CAS 2921 88 2. The Insecticide Resistance Action Committee IRAC classified Fipronil post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1B. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Fipronil for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.075 mg/L high exposure HE 0.3 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Fipronil CAS 120068 37 3 TraceCERT® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R843,SAMEA7678323,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678323|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9854:R843|age:96|broker name:ArrayExpress|cas:120068 37 3|common name:zebrafish|condition:control|developmental stage:larval stage|extraction date:27.01.2020|genotype:wild type genotype|nominal conc:0|organism part:whole organism|pooled individuals:10|rin:9.5|sample name:E MTAB 9854:R843|sex:mixed|strain:AB|tank spawning group:T10,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,E MTAB 9854:R843 s,R843 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Fipronil CAS 120068 37 3 TraceCERT® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125511,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Fipronil below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,R843.fastq.gz,fastq,1872733183.0,37120348.0,E MTAB 9854:R843,0:50.45 1:0,A:484141911;C:448431115;G:431926100;T:502374612;N:5859445,50,0,,,484141911,448431115,431926100,502374612,5859445,ERX4777538,ERS5435304,ERA3184565,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93265,,0.11236,,0.64585,,0.47631,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10102,ERR4910732,ERX4777555,ERS5435321,ERP125513,PRJEB41698,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,E-MTAB-9855,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos 96 hpf as aquatic vertebrate non target model exposed to sub lethal concentrations of the carbamate insecticide Carbaryl CAS 63 25 2. The Insecticide Resistance Action Committee IRAC classified Carbaryl post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1A. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Carbaryl for xxx hours under semi static conditions. Each test comprised of a low exposure LE 275 µg/L high exposure HE 1100 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Carbaryl CAS 63 25 2 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature.The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1201,SAMEA7678340,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678340|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9855:R1201|age:96|broker name:ArrayExpress|cas:63 25 2|common name:zebrafish|condition:low exposure|developmental stage:larval day 4|genotype:wild type genotype|nominal conc:0.000275|organism part:whole organism|pooled individuals:10|rin:8.9|sample name:E MTAB 9855:R1201|sex:mixed|strain:AB|tank spawning group:T19,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,E MTAB 9855:R1201 s,R1201 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Carbaryl CAS 63 25 2 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature.The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:carbaryl|Experimental Factor: dose:0.000275,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125513,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p915sR1201_sr.fastq.gz,fastq,1737913920.0,34408070.0,E MTAB 9855:R1201,0:50.51 1:0,A:443913578;C:424195235;G:405696397;T:462851439;N:1257271,50,0,,,443913578,424195235,405696397,462851439,1257271,ERX4777555,ERS5435321,ERA3184567,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93793,,0.0984,,0.65242,,0.48485,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10103,ERR4910731,ERX4777554,ERS5435320,ERP125513,PRJEB41698,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,E-MTAB-9855,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos 96 hpf as aquatic vertebrate non target model exposed to sub lethal concentrations of the carbamate insecticide Carbaryl CAS 63 25 2. The Insecticide Resistance Action Committee IRAC classified Carbaryl post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1A. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Carbaryl for xxx hours under semi static conditions. Each test comprised of a low exposure LE 275 µg/L high exposure HE 1100 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Carbaryl CAS 63 25 2 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature.The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1197,SAMEA7678339,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678339|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9855:R1197|age:96|broker name:ArrayExpress|cas:63 25 2|common name:zebrafish|condition:low exposure|developmental stage:larval day 4|genotype:wild type genotype|nominal conc:0.000275|organism part:whole organism|pooled individuals:10|rin:8.7|sample name:E MTAB 9855:R1197|sex:mixed|strain:AB|tank spawning group:T13,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,E MTAB 9855:R1197 s,R1197 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Carbaryl CAS 63 25 2 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature.The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:carbaryl|Experimental Factor: dose:0.000275,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125513,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p915sR1197_sr.fastq.gz,fastq,1882560681.0,37272626.0,E MTAB 9855:R1197,0:50.51 1:0,A:482382976;C:456835796;G:439735154;T:502548189;N:1058566,50,0,,,482382976,456835796,439735154,502548189,1058566,ERX4777554,ERS5435320,ERA3184567,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93062,,0.10062,,0.65005,,0.48156,,48,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10104,ERR4910730,ERX4777553,ERS5435319,ERP125513,PRJEB41698,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,E-MTAB-9855,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos 96 hpf as aquatic vertebrate non target model exposed to sub lethal concentrations of the carbamate insecticide Carbaryl CAS 63 25 2. The Insecticide Resistance Action Committee IRAC classified Carbaryl post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1A. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Carbaryl for xxx hours under semi static conditions. Each test comprised of a low exposure LE 275 µg/L high exposure HE 1100 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Carbaryl CAS 63 25 2 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature.The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1193,SAMEA7678338,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678338|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9855:R1193|age:96|broker name:ArrayExpress|cas:63 25 2|common name:zebrafish|condition:low exposure|developmental stage:larval day 4|genotype:wild type genotype|nominal conc:0.000275|organism part:whole organism|pooled individuals:10|rin:9.1|sample name:E MTAB 9855:R1193|sex:mixed|strain:AB|tank spawning group:T12,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,E MTAB 9855:R1193 s,R1193 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Carbaryl CAS 63 25 2 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature.The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:carbaryl|Experimental Factor: dose:0.000275,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125513,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p915sR1193_sr.fastq.gz,fastq,1645608286.0,32581218.0,E MTAB 9855:R1193,0:50.51 1:0,A:416942394;C:405832214;G:387058716;T:434382894;N:1392068,50,0,,,416942394,405832214,387058716,434382894,1392068,ERX4777553,ERS5435319,ERA3184567,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.94175,,0.10114,,0.65062,,0.48612,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10105,ERR4910729,ERX4777552,ERS5435318,ERP125513,PRJEB41698,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,E-MTAB-9855,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos 96 hpf as aquatic vertebrate non target model exposed to sub lethal concentrations of the carbamate insecticide Carbaryl CAS 63 25 2. The Insecticide Resistance Action Committee IRAC classified Carbaryl post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1A. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Carbaryl for xxx hours under semi static conditions. Each test comprised of a low exposure LE 275 µg/L high exposure HE 1100 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Carbaryl CAS 63 25 2 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature.The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1203,SAMEA7678337,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678337|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9855:R1203|age:96|broker name:ArrayExpress|cas:63 25 2|common name:zebrafish|condition:high exposure|developmental stage:larval day 4|genotype:wild type genotype|nominal conc:0.0011|organism part:whole organism|pooled individuals:10|rin:8.8|sample name:E MTAB 9855:R1203|sex:mixed|strain:AB|tank spawning group:T19,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,E MTAB 9855:R1203 s,R1203 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Carbaryl CAS 63 25 2 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature.The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:carbaryl|Experimental Factor: dose:0.0011,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125513,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p915sR1203_sr.fastq.gz,fastq,1538721898.0,30466055.0,E MTAB 9855:R1203,0:50.51 1:0,A:399907950;C:369324320;G:353100077;T:415526715;N:862836,50,0,,,399907950,369324320,353100077,415526715,862836,ERX4777552,ERS5435318,ERA3184567,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.92306,,0.10808,,0.64628,,0.48739,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10106,ERR4910728,ERX4777551,ERS5435317,ERP125513,PRJEB41698,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,E-MTAB-9855,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos 96 hpf as aquatic vertebrate non target model exposed to sub lethal concentrations of the carbamate insecticide Carbaryl CAS 63 25 2. The Insecticide Resistance Action Committee IRAC classified Carbaryl post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1A. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Carbaryl for xxx hours under semi static conditions. Each test comprised of a low exposure LE 275 µg/L high exposure HE 1100 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Carbaryl CAS 63 25 2 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature.The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1199,SAMEA7678336,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678336|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9855:R1199|age:96|broker name:ArrayExpress|cas:63 25 2|common name:zebrafish|condition:high exposure|developmental stage:larval day 4|genotype:wild type genotype|nominal conc:0.0011|organism part:whole organism|pooled individuals:10|rin:8.9|sample name:E MTAB 9855:R1199|sex:mixed|strain:AB|tank spawning group:T13,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,E MTAB 9855:R1199 s,R1199 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Carbaryl CAS 63 25 2 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature.The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:carbaryl|Experimental Factor: dose:0.0011,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125513,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p915sR1199_sr.fastq.gz,fastq,1574397484.0,31170945.0,E MTAB 9855:R1199,0:50.51 1:0,A:405653349;C:380978034;G:365558175;T:421378398;N:829528,50,0,,,405653349,380978034,365558175,421378398,829528,ERX4777551,ERS5435317,ERA3184567,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93022,,0.10311,,0.65115,,0.48604,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10107,ERR4910727,ERX4777550,ERS5435316,ERP125513,PRJEB41698,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,E-MTAB-9855,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos 96 hpf as aquatic vertebrate non target model exposed to sub lethal concentrations of the carbamate insecticide Carbaryl CAS 63 25 2. The Insecticide Resistance Action Committee IRAC classified Carbaryl post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1A. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Carbaryl for xxx hours under semi static conditions. Each test comprised of a low exposure LE 275 µg/L high exposure HE 1100 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Carbaryl CAS 63 25 2 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature.The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1195,SAMEA7678335,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678335|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9855:R1195|age:96|broker name:ArrayExpress|cas:63 25 2|common name:zebrafish|condition:high exposure|developmental stage:larval day 4|genotype:wild type genotype|nominal conc:0.0011|organism part:whole organism|pooled individuals:10|rin:8.8|sample name:E MTAB 9855:R1195|sex:mixed|strain:AB|tank spawning group:T12,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,E MTAB 9855:R1195 s,R1195 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Carbaryl CAS 63 25 2 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature.The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:carbaryl|Experimental Factor: dose:0.0011,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125513,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p915sR1195_sr.fastq.gz,fastq,2102149687.0,41635293.0,E MTAB 9855:R1195,0:50.49 1:0,A:541104841;C:508066558;G:486344627;T:563921691;N:2711970,50,0,,,541104841,508066558,486344627,563921691,2711970,ERX4777550,ERS5435316,ERA3184567,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93833,,0.11137,,0.64697,,0.4755,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10108,ERR4910726,ERX4777549,ERS5435315,ERP125513,PRJEB41698,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,E-MTAB-9855,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos 96 hpf as aquatic vertebrate non target model exposed to sub lethal concentrations of the carbamate insecticide Carbaryl CAS 63 25 2. The Insecticide Resistance Action Committee IRAC classified Carbaryl post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1A. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Carbaryl for xxx hours under semi static conditions. Each test comprised of a low exposure LE 275 µg/L high exposure HE 1100 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Carbaryl CAS 63 25 2 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature.The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1200,SAMEA7678334,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678334|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9855:R1200|age:96|broker name:ArrayExpress|cas:63 25 2|common name:zebrafish|condition:control|developmental stage:larval day 4|genotype:wild type genotype|nominal conc:0|organism part:whole organism|pooled individuals:10|rin:8.7|sample name:E MTAB 9855:R1200|sex:mixed|strain:AB|tank spawning group:T19,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,E MTAB 9855:R1200 s,R1200 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Carbaryl CAS 63 25 2 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature.The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125513,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p915sR1200_sr.fastq.gz,fastq,2998689981.0,59376762.0,E MTAB 9855:R1200,0:50.50 1:0,A:770180835;C:728231589;G:694302138;T:803817388;N:2158031,50,0,,,770180835,728231589,694302138,803817388,2158031,ERX4777549,ERS5435315,ERA3184567,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.92959,,0.10236,,0.64962,,0.47823,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10109,ERR4910725,ERX4777548,ERS5435314,ERP125513,PRJEB41698,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,E-MTAB-9855,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos 96 hpf as aquatic vertebrate non target model exposed to sub lethal concentrations of the carbamate insecticide Carbaryl CAS 63 25 2. The Insecticide Resistance Action Committee IRAC classified Carbaryl post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1A. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Carbaryl for xxx hours under semi static conditions. Each test comprised of a low exposure LE 275 µg/L high exposure HE 1100 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Carbaryl CAS 63 25 2 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature.The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1196,SAMEA7678333,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678333|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9855:R1196|age:96|broker name:ArrayExpress|cas:63 25 2|common name:zebrafish|condition:control|developmental stage:larval day 4|genotype:wild type genotype|nominal conc:0|organism part:whole organism|pooled individuals:10|rin:9|sample name:E MTAB 9855:R1196|sex:mixed|strain:AB|tank spawning group:T13,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,E MTAB 9855:R1196 s,R1196 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Carbaryl CAS 63 25 2 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature.The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125513,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p915sR1196_sr.fastq.gz,fastq,1580346161.0,31296644.0,E MTAB 9855:R1196,0:50.50 1:0,A:405887724;C:382519850;G:367322304;T:422929756;N:1686527,50,0,,,405887724,382519850,367322304,422929756,1686527,ERX4777548,ERS5435314,ERA3184567,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.92603,,0.10135,,0.64975,,0.47346,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10110,ERR4910724,ERX4777547,ERS5435313,ERP125513,PRJEB41698,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,E-MTAB-9855,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos 96 hpf as aquatic vertebrate non target model exposed to sub lethal concentrations of the carbamate insecticide Carbaryl CAS 63 25 2. The Insecticide Resistance Action Committee IRAC classified Carbaryl post its mode of action MoA in the target organism as an acetylcholinesterase AChE inhibitor Group 1A. The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Carbaryl for xxx hours under semi static conditions. Each test comprised of a low exposure LE 275 µg/L high exposure HE 1100 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Carbaryl CAS 63 25 2 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature.The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1192,SAMEA7678332,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7678332|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:29Z|INSDC last update:2021 10 15T00:15:29Z|INSDC status:public|Submitter Id:E MTAB 9855:R1192|age:96|broker name:ArrayExpress|cas:63 25 2|common name:zebrafish|condition:control|developmental stage:larval day 4|genotype:wild type genotype|nominal conc:0|organism part:whole organism|pooled individuals:10|rin:8.7|sample name:E MTAB 9855:R1192|sex:mixed|strain:AB|tank spawning group:T12,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,E MTAB 9855:R1192 s,R1192 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Carbaryl CAS 63 25 2 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature.The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125513,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Carbaryl below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p915sR1192_sr.fastq.gz,fastq,1330588291.0,26350032.0,E MTAB 9855:R1192,0:50.50 1:0,A:342926562;C:321435886;G:307380318;T:357452217;N:1393308,50,0,,,342926562,321435886,307380318,357452217,1393308,ERX4777547,ERS5435313,ERA3184567,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93927,,0.11145,,0.64329,,0.48211,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10117,ERR4968945,ERX4788142,ERS5455107,ERP125626,PRJEB41791,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E-MTAB-9859,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used neonicotinoid insecticide Imidacloprid CAS 138261 41 3. The Insecticide Resistance Action Committee IRAC classified Imidacloprid post its mode of action MoA in the target organism as a nicotinic acetylcholine receptor nAChR competitive modulator Group 4A The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Imidacloprid for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.015 mg/L medium exposure ME 0.03 mg/L high exposure HE 0.06 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R286,SAMEA7698592,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698592|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:30Z|INSDC last update:2021 10 15T00:15:30Z|INSDC status:public|Submitter Id:E MTAB 9859:R286|age:96|broker name:ArrayExpress|cas:138261 41 3|common name:zebrafish|condition:HighExposure|developmental stage:larval day 4|extraction date:18.03.2019|genotype:wild type genotype|nominal concentration:0 06|organism part:whole larvae|pooled individuals:10|rin:9 7|sample name:E MTAB 9859:R286|sex:mal & femal pooled|strain:AB|tank spawning group:T13 1,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E MTAB 9859:R286 s,R286 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:HighExposure|Experimental Factor: nominal concentration:0 06|Experimental Factor: dose:0 06|Experimental Factor: compound:imidacloprid,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125626,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p689sR286_sr.fastq.gz,fastq,1237910329.0,24520799.0,E MTAB 9859:R286,0:50.48 1:0,A:316319865;C:303280694;G:290244904;T:326544610;N:1520256,50,0,,,316319865,303280694,290244904,326544610,1520256,ERX4788142,ERS5455107,ERA3190564,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.94066,,0.10761,,0.6578,,0.46552,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10118,ERR4968944,ERX4788141,ERS5455106,ERP125626,PRJEB41791,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E-MTAB-9859,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used neonicotinoid insecticide Imidacloprid CAS 138261 41 3. The Insecticide Resistance Action Committee IRAC classified Imidacloprid post its mode of action MoA in the target organism as a nicotinic acetylcholine receptor nAChR competitive modulator Group 4A The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Imidacloprid for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.015 mg/L medium exposure ME 0.03 mg/L high exposure HE 0.06 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R275,SAMEA7698591,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698591|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:30Z|INSDC last update:2021 10 15T00:15:30Z|INSDC status:public|Submitter Id:E MTAB 9859:R275|age:96|broker name:ArrayExpress|cas:138261 41 3|common name:zebrafish|condition:HighExposure|developmental stage:larval day 4|extraction date:22.02.2019|genotype:wild type genotype|nominal concentration:0 06|organism part:whole larvae|pooled individuals:10|rin:9 8|sample name:E MTAB 9859:R275|sex:mal & femal pooled|strain:AB|tank spawning group:T8,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E MTAB 9859:R275 s,R275 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:HighExposure|Experimental Factor: nominal concentration:0 06|Experimental Factor: dose:0 06|Experimental Factor: compound:imidacloprid,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125626,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p689sR275_sr.fastq.gz,fastq,1530123976.0,30313143.0,E MTAB 9859:R275,0:50.48 1:0,A:391068440;C:369486566;G:359123887;T:407522026;N:2923057,50,0,,,391068440,369486566,359123887,407522026,2923057,ERX4788141,ERS5455106,ERA3190564,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.94058,,0.1128,,0.65997,,0.46991,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10119,ERR4968943,ERX4788140,ERS5455105,ERP125626,PRJEB41791,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E-MTAB-9859,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used neonicotinoid insecticide Imidacloprid CAS 138261 41 3. The Insecticide Resistance Action Committee IRAC classified Imidacloprid post its mode of action MoA in the target organism as a nicotinic acetylcholine receptor nAChR competitive modulator Group 4A The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Imidacloprid for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.015 mg/L medium exposure ME 0.03 mg/L high exposure HE 0.06 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R268,SAMEA7698590,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698590|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:30Z|INSDC last update:2021 10 15T00:15:30Z|INSDC status:public|Submitter Id:E MTAB 9859:R268|age:96|broker name:ArrayExpress|cas:138261 41 3|common name:zebrafish|condition:HighExposure|developmental stage:larval day 4|extraction date:22.02.2019|genotype:wild type genotype|nominal concentration:0 06|organism part:whole larvae|pooled individuals:10|rin:10|sample name:E MTAB 9859:R268|sex:mal & femal pooled|strain:AB|tank spawning group:T14 3,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E MTAB 9859:R268 s,R268 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:HighExposure|Experimental Factor: nominal concentration:0 06|Experimental Factor: dose:0 06|Experimental Factor: compound:imidacloprid,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125626,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p689sR268_sr.fastq.gz,fastq,1185384697.0,23473359.0,E MTAB 9859:R268,0:50.50 1:0,A:305557028;C:287664002;G:275082416;T:316155443;N:925808,50,0,,,305557028,287664002,275082416,316155443,925808,ERX4788140,ERS5455105,ERA3190564,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93951,,0.11512,,0.65896,,0.47402,,48,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10120,ERR4968942,ERX4788139,ERS5455104,ERP125626,PRJEB41791,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E-MTAB-9859,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used neonicotinoid insecticide Imidacloprid CAS 138261 41 3. The Insecticide Resistance Action Committee IRAC classified Imidacloprid post its mode of action MoA in the target organism as a nicotinic acetylcholine receptor nAChR competitive modulator Group 4A The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Imidacloprid for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.015 mg/L medium exposure ME 0.03 mg/L high exposure HE 0.06 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R285,SAMEA7698589,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698589|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:30Z|INSDC last update:2021 10 15T00:15:30Z|INSDC status:public|Submitter Id:E MTAB 9859:R285|age:96|broker name:ArrayExpress|cas:138261 41 3|common name:zebrafish|condition:MidExposure|developmental stage:larval day 4|extraction date:18.03.2019|genotype:wild type genotype|nominal concentration:0 03|organism part:whole larvae|pooled individuals:10|rin:9 6|sample name:E MTAB 9859:R285|sex:mal & femal pooled|strain:AB|tank spawning group:T13 1,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E MTAB 9859:R285 s,R285 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:MidExposure|Experimental Factor: nominal concentration:0 03|Experimental Factor: dose:0 03|Experimental Factor: compound:imidacloprid,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125626,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p823sR285_sr.fastq.gz,fastq,1237013890.0,24497402.0,E MTAB 9859:R285,0:50.50 1:0,A:318805223;C:298918575;G:287178284;T:330710282;N:1401526,50,0,,,318805223,298918575,287178284,330710282,1401526,ERX4788139,ERS5455104,ERA3190564,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93827,,0.10255,,0.64993,,0.47369,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10121,ERR4968941,ERX4788138,ERS5455103,ERP125626,PRJEB41791,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E-MTAB-9859,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used neonicotinoid insecticide Imidacloprid CAS 138261 41 3. The Insecticide Resistance Action Committee IRAC classified Imidacloprid post its mode of action MoA in the target organism as a nicotinic acetylcholine receptor nAChR competitive modulator Group 4A The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Imidacloprid for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.015 mg/L medium exposure ME 0.03 mg/L high exposure HE 0.06 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R273,SAMEA7698588,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698588|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:30Z|INSDC last update:2021 10 15T00:15:30Z|INSDC status:public|Submitter Id:E MTAB 9859:R273|age:96|broker name:ArrayExpress|cas:138261 41 3|common name:zebrafish|condition:MidExposure|developmental stage:larval day 4|extraction date:22.02.2019|genotype:wild type genotype|nominal concentration:0 03|organism part:whole larvae|pooled individuals:10|rin:10|sample name:E MTAB 9859:R273|sex:mal & femal pooled|strain:AB|tank spawning group:T8,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E MTAB 9859:R273 s,R273 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:MidExposure|Experimental Factor: nominal concentration:0 03|Experimental Factor: dose:0 03|Experimental Factor: compound:imidacloprid,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125626,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p823sR273_sr.fastq.gz,fastq,1223214587.0,24228262.0,E MTAB 9859:R273,0:50.49 1:0,A:317013129;C:293403474;G:282025942;T:328880126;N:1891916,50,0,,,317013129,293403474,282025942,328880126,1891916,ERX4788138,ERS5455103,ERA3190564,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93783,,0.10916,,0.65242,,0.47563,,49,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10122,ERR4968940,ERX4788137,ERS5455102,ERP125626,PRJEB41791,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E-MTAB-9859,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used neonicotinoid insecticide Imidacloprid CAS 138261 41 3. The Insecticide Resistance Action Committee IRAC classified Imidacloprid post its mode of action MoA in the target organism as a nicotinic acetylcholine receptor nAChR competitive modulator Group 4A The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Imidacloprid for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.015 mg/L medium exposure ME 0.03 mg/L high exposure HE 0.06 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R267,SAMEA7698587,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698587|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:30Z|INSDC last update:2021 10 15T00:15:30Z|INSDC status:public|Submitter Id:E MTAB 9859:R267|age:96|broker name:ArrayExpress|cas:138261 41 3|common name:zebrafish|condition:MidExposure|developmental stage:larval day 4|extraction date:22.02.2019|genotype:wild type genotype|nominal concentration:0 03|organism part:whole larvae|pooled individuals:10|rin:10|sample name:E MTAB 9859:R267|sex:mal & femal pooled|strain:AB|tank spawning group:T14 3,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E MTAB 9859:R267 s,R267 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:MidExposure|Experimental Factor: nominal concentration:0 03|Experimental Factor: dose:0 03|Experimental Factor: compound:imidacloprid,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125626,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p823sR267_sr.fastq.gz,fastq,1507738144.0,29878977.0,E MTAB 9859:R267,0:50.46 1:0,A:389888230;C:360392926;G:348869994;T:404349593;N:4237401,50,0,,,389888230,360392926,348869994,404349593,4237401,ERX4788137,ERS5455102,ERA3190564,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.94014,,0.11127,,0.65066,,0.47853,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10123,ERR4968939,ERX4788136,ERS5455101,ERP125626,PRJEB41791,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E-MTAB-9859,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used neonicotinoid insecticide Imidacloprid CAS 138261 41 3. The Insecticide Resistance Action Committee IRAC classified Imidacloprid post its mode of action MoA in the target organism as a nicotinic acetylcholine receptor nAChR competitive modulator Group 4A The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Imidacloprid for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.015 mg/L medium exposure ME 0.03 mg/L high exposure HE 0.06 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R284,SAMEA7698586,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698586|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:30Z|INSDC last update:2021 10 15T00:15:30Z|INSDC status:public|Submitter Id:E MTAB 9859:R284|age:96|broker name:ArrayExpress|cas:138261 41 3|common name:zebrafish|condition:LowExposure|developmental stage:larval day 4|extraction date:18.03.2019|genotype:wild type genotype|nominal concentration:15|organism part:whole larvae|pooled individuals:10|rin:9 6|sample name:E MTAB 9859:R284|sex:mal & femal pooled|strain:AB|tank spawning group:T13 1,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E MTAB 9859:R284 s,R284 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:LowExposure|Experimental Factor: nominal concentration:15|Experimental Factor: dose:15|Experimental Factor: compound:imidacloprid,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125626,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p689sR284_sr.fastq.gz,fastq,1278659450.0,25321525.0,E MTAB 9859:R284,0:50.50 1:0,A:328307749;C:311381078;G:298180080;T:339736744;N:1053799,50,0,,,328307749,311381078,298180080,339736744,1053799,ERX4788136,ERS5455101,ERA3190564,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93931,,0.10051,,0.65316,,0.47695,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10124,ERR4968938,ERX4788135,ERS5455100,ERP125626,PRJEB41791,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E-MTAB-9859,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used neonicotinoid insecticide Imidacloprid CAS 138261 41 3. The Insecticide Resistance Action Committee IRAC classified Imidacloprid post its mode of action MoA in the target organism as a nicotinic acetylcholine receptor nAChR competitive modulator Group 4A The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Imidacloprid for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.015 mg/L medium exposure ME 0.03 mg/L high exposure HE 0.06 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R272,SAMEA7698585,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698585|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:30Z|INSDC last update:2021 10 15T00:15:30Z|INSDC status:public|Submitter Id:E MTAB 9859:R272|age:96|broker name:ArrayExpress|cas:138261 41 3|common name:zebrafish|condition:LowExposure|developmental stage:larval day 4|extraction date:22.02.2019|genotype:wild type genotype|nominal concentration:15|organism part:whole larvae|pooled individuals:10|rin:10|sample name:E MTAB 9859:R272|sex:mal & femal pooled|strain:AB|tank spawning group:T8,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E MTAB 9859:R272 s,R272 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:LowExposure|Experimental Factor: nominal concentration:15|Experimental Factor: dose:15|Experimental Factor: compound:imidacloprid,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125626,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p689sR272_sr.fastq.gz,fastq,1444590100.0,28607531.0,E MTAB 9859:R272,0:50.50 1:0,A:372710449;C:349880176;G:334911408;T:385848878;N:1239189,50,0,,,372710449,349880176,334911408,385848878,1239189,ERX4788135,ERS5455100,ERA3190564,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93838,,0.10963,,0.65178,,0.47899,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10125,ERR4968937,ERX4788134,ERS5455099,ERP125626,PRJEB41791,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E-MTAB-9859,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used neonicotinoid insecticide Imidacloprid CAS 138261 41 3. The Insecticide Resistance Action Committee IRAC classified Imidacloprid post its mode of action MoA in the target organism as a nicotinic acetylcholine receptor nAChR competitive modulator Group 4A The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Imidacloprid for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.015 mg/L medium exposure ME 0.03 mg/L high exposure HE 0.06 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R266,SAMEA7698584,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698584|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:30Z|INSDC last update:2021 10 15T00:15:30Z|INSDC status:public|Submitter Id:E MTAB 9859:R266|age:96|broker name:ArrayExpress|cas:138261 41 3|common name:zebrafish|condition:LowExposure|developmental stage:larval day 4|extraction date:22.02.2019|genotype:wild type genotype|nominal concentration:15|organism part:whole larvae|pooled individuals:10|rin:10|sample name:E MTAB 9859:R266|sex:mal & femal pooled|strain:AB|tank spawning group:T14 3,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E MTAB 9859:R266 s,R266 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:LowExposure|Experimental Factor: nominal concentration:15|Experimental Factor: dose:15|Experimental Factor: compound:imidacloprid,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125626,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p689sR266_sr.fastq.gz,fastq,1061121934.0,21013546.0,E MTAB 9859:R266,0:50.50 1:0,A:276035557;C:255134233;G:243420790;T:285689121;N:842233,50,0,,,276035557,255134233,243420790,285689121,842233,ERX4788134,ERS5455099,ERA3190564,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93605,,0.11404,,0.65281,,0.48179,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10126,ERR4968936,ERX4788133,ERS5455098,ERP125626,PRJEB41791,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E-MTAB-9859,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used neonicotinoid insecticide Imidacloprid CAS 138261 41 3. The Insecticide Resistance Action Committee IRAC classified Imidacloprid post its mode of action MoA in the target organism as a nicotinic acetylcholine receptor nAChR competitive modulator Group 4A The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Imidacloprid for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.015 mg/L medium exposure ME 0.03 mg/L high exposure HE 0.06 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R283,SAMEA7698583,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698583|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:30Z|INSDC last update:2021 10 15T00:15:30Z|INSDC status:public|Submitter Id:E MTAB 9859:R283|age:96|broker name:ArrayExpress|cas:138261 41 3|common name:zebrafish|condition:Control|developmental stage:larval day 4|extraction date:18.03.2019|genotype:wild type genotype|nominal concentration:0|organism part:whole larvae|pooled individuals:10|rin:9 7|sample name:E MTAB 9859:R283|sex:mal & femal pooled|strain:AB|tank spawning group:T13 1,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E MTAB 9859:R283 s,R283 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:Control|Experimental Factor: nominal concentration:0|Experimental Factor: dose:0|Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125626,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p689sR283_sr.fastq.gz,fastq,1330144467.0,26345369.0,E MTAB 9859:R283,0:50.49 1:0,A:343869074;C:322612240;G:306958987;T:355387615;N:1316551,50,0,,,343869074,322612240,306958987,355387615,1316551,ERX4788133,ERS5455098,ERA3190564,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93681,,0.10977,,0.65311,,0.48251,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10127,ERR4968935,ERX4788132,ERS5455097,ERP125626,PRJEB41791,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E-MTAB-9859,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used neonicotinoid insecticide Imidacloprid CAS 138261 41 3. The Insecticide Resistance Action Committee IRAC classified Imidacloprid post its mode of action MoA in the target organism as a nicotinic acetylcholine receptor nAChR competitive modulator Group 4A The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Imidacloprid for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.015 mg/L medium exposure ME 0.03 mg/L high exposure HE 0.06 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R271,SAMEA7698582,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698582|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:30Z|INSDC last update:2021 10 15T00:15:30Z|INSDC status:public|Submitter Id:E MTAB 9859:R271|age:96|broker name:ArrayExpress|cas:138261 41 3|common name:zebrafish|condition:Control|developmental stage:larval day 4|extraction date:22.02.2019|genotype:wild type genotype|nominal concentration:0|organism part:whole larvae|pooled individuals:10|rin:10|sample name:E MTAB 9859:R271|sex:mal & femal pooled|strain:AB|tank spawning group:T8,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E MTAB 9859:R271 s,R271 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:Control|Experimental Factor: nominal concentration:0|Experimental Factor: dose:0|Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125626,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p689sR271_sr.fastq.gz,fastq,1184008393.0,23449500.0,E MTAB 9859:R271,0:50.49 1:0,A:305676242;C:287310817;G:274322930;T:315613198;N:1085206,50,0,,,305676242,287310817,274322930,315613198,1085206,ERX4788132,ERS5455097,ERA3190564,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93602,,0.10767,,0.6547,,0.46846,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10128,ERR4968934,ERX4788131,ERS5455096,ERP125626,PRJEB41791,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E-MTAB-9859,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of the heavily used neonicotinoid insecticide Imidacloprid CAS 138261 41 3. The Insecticide Resistance Action Committee IRAC classified Imidacloprid post its mode of action MoA in the target organism as a nicotinic acetylcholine receptor nAChR competitive modulator Group 4A The goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Imidacloprid for xxx hours under semi static conditions. Each test comprised of a low exposure LE 0.015 mg/L medium exposure ME 0.03 mg/L high exposure HE 0.06 mg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for data normalization and differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R265,SAMEA7698581,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698581|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:30Z|INSDC last update:2021 10 15T00:15:30Z|INSDC status:public|Submitter Id:E MTAB 9859:R265|age:96|broker name:ArrayExpress|cas:138261 41 3|common name:zebrafish|condition:Control|developmental stage:larval day 4|extraction date:22.02.2019|genotype:wild type genotype|nominal concentration:0|organism part:whole larvae|pooled individuals:10|rin:10|sample name:E MTAB 9859:R265|sex:mal & femal pooled|strain:AB|tank spawning group:T14 3,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,E MTAB 9859:R265 s,R265 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Imidacloprid CAS 138261 41 3 PESTANAL® analytical standard purity ≥ 98.0 % was purchased from Merck KGgA Darmstadt Germany. Test solutions were freshly prepared one day prior starting the exposure experiment. For that the solutions were setup in 200 ml of Cu reduced water and stirred in the dark for 3 hours at room temperature before adding 8 ml of the respective test solution to the glass wells for pre saturation overnight. The remaining test solutions were stored in the dark at room temperature and the solution in the well was renewed the next day with the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:Control|Experimental Factor: nominal concentration:0|Experimental Factor: dose:0|Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125626,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of Imidacloprid below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p689sR265_sr.fastq.gz,fastq,1305790358.0,25859113.0,E MTAB 9859:R265,0:50.50 1:0,A:338087552;C:316509262;G:301805722;T:348360089;N:1027733,50,0,,,338087552,316509262,301805722,348360089,1027733,ERX4788131,ERS5455096,ERA3190564,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93665,,0.10959,,0.65411,,0.48771,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10129,ERR5017657,ERX4826558,ERS5489438,ERP125627,PRJEB41792,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E-MTAB-9860,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of synthetic organochloride insecticide Methoxychlor CAS 72 43 5. It is structurally highly similar to its precursor molecule DDT Dichlorodiphenyltrichloroethane. Like its precursor today Methoxychlor is banned from the use as pesticide in the United States and the European Union due to its acute toxicity high bioaccumulation potential and endocrine disruption activity. The Insecticide Resistance Action Committee IRAC classified Methoxychlor post its mode of action MoA in the target organism as a sodium channel modulator Group 3B. Our goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms for this substance of particularly high environmental concern. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Methoxychlor for xxx hours under semi static conditions. Each test comprised of a low exposure LE 20 µg/L high exposure HE 180 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R969,SAMEA7742085,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7742085|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:31Z|INSDC last update:2021 10 15T00:15:31Z|INSDC status:public|Submitter Id:E MTAB 98601608543770:R969|age:96|broker name:ArrayExpress|cas:72 43 5|common name:zebrafish|condition:MidExposure|developmental stage:larval day 4|extraction date:17.02.2020|genotype:wild type genotype|nominal conc:0 00006|organism part:whole larvae|pooled individuals:10|rin:9 3|sample name:E MTAB 98601608543770:R969|sex:male & female pooled|strain:AB|tank spawning group:T11,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E MTAB 98601608543770:R969 s,R969 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:MidExposure|Experimental Factor: compound:methoxychlor|Experimental Factor: dose:0 00006|Experimental Factor: nominal conc:0 00006,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125627,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p946sR969_sr.fastq.gz,fastq,1298525482.0,25717800.0,E MTAB 98601608543770:R969,0:50.49 1:0,A:337612374;C:313888872;G:298044250;T:347704703;N:1275283,50,0,,,337612374,313888872,298044250,347704703,1275283,ERX4826558,ERS5489438,ERA3202392,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.932,,0.11102,,0.65157,,0.47488,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10130,ERR5017656,ERX4826557,ERS5489437,ERP125627,PRJEB41792,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E-MTAB-9860,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of synthetic organochloride insecticide Methoxychlor CAS 72 43 5. It is structurally highly similar to its precursor molecule DDT Dichlorodiphenyltrichloroethane. Like its precursor today Methoxychlor is banned from the use as pesticide in the United States and the European Union due to its acute toxicity high bioaccumulation potential and endocrine disruption activity. The Insecticide Resistance Action Committee IRAC classified Methoxychlor post its mode of action MoA in the target organism as a sodium channel modulator Group 3B. Our goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms for this substance of particularly high environmental concern. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Methoxychlor for xxx hours under semi static conditions. Each test comprised of a low exposure LE 20 µg/L high exposure HE 180 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R965,SAMEA7742084,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7742084|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:31Z|INSDC last update:2021 10 15T00:15:31Z|INSDC status:public|Submitter Id:E MTAB 98601608543770:R965|age:96|broker name:ArrayExpress|cas:72 43 5|common name:zebrafish|condition:MidExposure|developmental stage:larval day 4|extraction date:17.02.2020|genotype:wild type genotype|nominal conc:0 00006|organism part:whole larvae|pooled individuals:10|rin:8 9|sample name:E MTAB 98601608543770:R965|sex:male & female pooled|strain:AB|tank spawning group:T10,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E MTAB 98601608543770:R965 s,R965 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:MidExposure|Experimental Factor: compound:methoxychlor|Experimental Factor: dose:0 00006|Experimental Factor: nominal conc:0 00006,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125627,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p946sR965_sr.fastq.gz,fastq,1934920385.0,38320190.0,E MTAB 98601608543770:R965,0:50.49 1:0,A:498112571;C:469867864;G:448829396;T:516385769;N:1724785,50,0,,,498112571,469867864,448829396,516385769,1724785,ERX4826557,ERS5489437,ERA3202392,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93881,,0.1002,,0.6464,,0.47456,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10131,ERR5017655,ERX4826556,ERS5489436,ERP125627,PRJEB41792,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E-MTAB-9860,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of synthetic organochloride insecticide Methoxychlor CAS 72 43 5. It is structurally highly similar to its precursor molecule DDT Dichlorodiphenyltrichloroethane. Like its precursor today Methoxychlor is banned from the use as pesticide in the United States and the European Union due to its acute toxicity high bioaccumulation potential and endocrine disruption activity. The Insecticide Resistance Action Committee IRAC classified Methoxychlor post its mode of action MoA in the target organism as a sodium channel modulator Group 3B. Our goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms for this substance of particularly high environmental concern. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Methoxychlor for xxx hours under semi static conditions. Each test comprised of a low exposure LE 20 µg/L high exposure HE 180 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R961,SAMEA7742083,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7742083|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:31Z|INSDC last update:2021 10 15T00:15:31Z|INSDC status:public|Submitter Id:E MTAB 98601608543770:R961|age:96|broker name:ArrayExpress|cas:72 43 5|common name:zebrafish|condition:MidExposure|developmental stage:larval day 4|extraction date:17.02.2020|genotype:wild type genotype|nominal conc:0 00006|organism part:whole larvae|pooled individuals:10|rin:9 2|sample name:E MTAB 98601608543770:R961|sex:male & female pooled|strain:AB|tank spawning group:T7,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E MTAB 98601608543770:R961 s,R961 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:MidExposure|Experimental Factor: compound:methoxychlor|Experimental Factor: dose:0 00006|Experimental Factor: nominal conc:0 00006,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125627,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p946sR961_sr.fastq.gz,fastq,1545220277.0,30598922.0,E MTAB 98601608543770:R961,0:50.50 1:0,A:394686821;C:378912771;G:361041863;T:409273024;N:1305798,50,0,,,394686821,378912771,361041863,409273024,1305798,ERX4826556,ERS5489436,ERA3202392,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.94085,,0.09116,,0.65007,,0.47771,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10132,ERR4968954,ERX4788151,ERS5455116,ERP125627,PRJEB41792,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E-MTAB-9860,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of synthetic organochloride insecticide Methoxychlor CAS 72 43 5. It is structurally highly similar to its precursor molecule DDT Dichlorodiphenyltrichloroethane. Like its precursor today Methoxychlor is banned from the use as pesticide in the United States and the European Union due to its acute toxicity high bioaccumulation potential and endocrine disruption activity. The Insecticide Resistance Action Committee IRAC classified Methoxychlor post its mode of action MoA in the target organism as a sodium channel modulator Group 3B. Our goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms for this substance of particularly high environmental concern. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Methoxychlor for xxx hours under semi static conditions. Each test comprised of a low exposure LE 20 µg/L high exposure HE 180 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R970,SAMEA7698601,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698601|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:31Z|INSDC last update:2021 10 15T00:15:31Z|INSDC status:public|Submitter Id:E MTAB 9860:R970|age:96|broker name:ArrayExpress|cas:72 43 5|common name:zebrafish|condition:HighExposure|developmental stage:larval day 4|extraction date:17.02.2020|genotype:wild type genotype|nominal conc:0 00018|organism part:whole larvae|pooled individuals:10|rin:8 4|sample name:E MTAB 9860:R970|sex:male & female pooled|strain:AB|tank spawning group:T11,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E MTAB 9860:R970 s,R970 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:HighExposure|Experimental Factor: dose:0 00018|Experimental Factor: nominal conc:0 00018|Experimental Factor: compound:methoxychlor,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125627,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p915sR970_sr.fastq.gz,fastq,1450580646.0,28739452.0,E MTAB 9860:R970,0:50.47 1:0,A:373203876;C:350529390;G:338021223;T:386562361;N:2263796,50,0,,,373203876,350529390,338021223,386562361,2263796,ERX4788151,ERS5455116,ERA3190565,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.9376,,0.10301,,0.65287,,0.47862,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10133,ERR4968953,ERX4788150,ERS5455115,ERP125627,PRJEB41792,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E-MTAB-9860,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of synthetic organochloride insecticide Methoxychlor CAS 72 43 5. It is structurally highly similar to its precursor molecule DDT Dichlorodiphenyltrichloroethane. Like its precursor today Methoxychlor is banned from the use as pesticide in the United States and the European Union due to its acute toxicity high bioaccumulation potential and endocrine disruption activity. The Insecticide Resistance Action Committee IRAC classified Methoxychlor post its mode of action MoA in the target organism as a sodium channel modulator Group 3B. Our goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms for this substance of particularly high environmental concern. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Methoxychlor for xxx hours under semi static conditions. Each test comprised of a low exposure LE 20 µg/L high exposure HE 180 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R966,SAMEA7698600,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698600|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:31Z|INSDC last update:2021 10 15T00:15:31Z|INSDC status:public|Submitter Id:E MTAB 9860:R966|age:96|broker name:ArrayExpress|cas:72 43 5|common name:zebrafish|condition:HighExposure|developmental stage:larval day 4|extraction date:17.02.2020|genotype:wild type genotype|nominal conc:0 00018|organism part:whole larvae|pooled individuals:10|rin:8 9|sample name:E MTAB 9860:R966|sex:male & female pooled|strain:AB|tank spawning group:T10,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E MTAB 9860:R966 s,R966 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:HighExposure|Experimental Factor: dose:0 00018|Experimental Factor: nominal conc:0 00018|Experimental Factor: compound:methoxychlor,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125627,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p915sR966_sr.fastq.gz,fastq,1567145939.0,31054634.0,E MTAB 9860:R966,0:50.46 1:0,A:401414524;C:379485206;G:367308924;T:415939815;N:2997470,50,0,,,401414524,379485206,367308924,415939815,2997470,ERX4788150,ERS5455115,ERA3190565,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93615,,0.10572,,0.65176,,0.48206,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10134,ERR4968952,ERX4788149,ERS5455114,ERP125627,PRJEB41792,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E-MTAB-9860,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of synthetic organochloride insecticide Methoxychlor CAS 72 43 5. It is structurally highly similar to its precursor molecule DDT Dichlorodiphenyltrichloroethane. Like its precursor today Methoxychlor is banned from the use as pesticide in the United States and the European Union due to its acute toxicity high bioaccumulation potential and endocrine disruption activity. The Insecticide Resistance Action Committee IRAC classified Methoxychlor post its mode of action MoA in the target organism as a sodium channel modulator Group 3B. Our goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms for this substance of particularly high environmental concern. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Methoxychlor for xxx hours under semi static conditions. Each test comprised of a low exposure LE 20 µg/L high exposure HE 180 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R962,SAMEA7698599,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698599|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:31Z|INSDC last update:2021 10 15T00:15:31Z|INSDC status:public|Submitter Id:E MTAB 9860:R962|age:96|broker name:ArrayExpress|cas:72 43 5|common name:zebrafish|condition:HighExposure|developmental stage:larval day 4|extraction date:17.02.2020|genotype:wild type genotype|nominal conc:0 00018|organism part:whole larvae|pooled individuals:10|rin:9 4|sample name:E MTAB 9860:R962|sex:male & female pooled|strain:AB|tank spawning group:T7,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E MTAB 9860:R962 s,R962 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:HighExposure|Experimental Factor: dose:0 00018|Experimental Factor: nominal conc:0 00018|Experimental Factor: compound:methoxychlor,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125627,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p915sR962_sr.fastq.gz,fastq,1905870912.0,37761427.0,E MTAB 9860:R962,0:50.47 1:0,A:486923070;C:463730576;G:447319499;T:504586593;N:3311174,50,0,,,486923070,463730576,447319499,504586593,3311174,ERX4788149,ERS5455114,ERA3190565,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93804,,0.09568,,0.65368,,0.47176,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10135,ERR4968951,ERX4788148,ERS5455113,ERP125627,PRJEB41792,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E-MTAB-9860,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of synthetic organochloride insecticide Methoxychlor CAS 72 43 5. It is structurally highly similar to its precursor molecule DDT Dichlorodiphenyltrichloroethane. Like its precursor today Methoxychlor is banned from the use as pesticide in the United States and the European Union due to its acute toxicity high bioaccumulation potential and endocrine disruption activity. The Insecticide Resistance Action Committee IRAC classified Methoxychlor post its mode of action MoA in the target organism as a sodium channel modulator Group 3B. Our goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms for this substance of particularly high environmental concern. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Methoxychlor for xxx hours under semi static conditions. Each test comprised of a low exposure LE 20 µg/L high exposure HE 180 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R968,SAMEA7698598,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698598|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:31Z|INSDC last update:2021 10 15T00:15:31Z|INSDC status:public|Submitter Id:E MTAB 9860:R968|age:96|broker name:ArrayExpress|cas:72 43 5|common name:zebrafish|condition:LowExposure|developmental stage:larval day 4|extraction date:17.02.2020|genotype:wild type genotype|nominal conc:0 00002|organism part:whole larvae|pooled individuals:10|rin:8 5|sample name:E MTAB 9860:R968|sex:male & female pooled|strain:AB|tank spawning group:T11,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E MTAB 9860:R968 s,R968 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:LowExposure|Experimental Factor: dose:0 00002|Experimental Factor: nominal conc:0 00002|Experimental Factor: compound:methoxychlor,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125627,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p915sR968_sr.fastq.gz,fastq,1734649737.0,34362680.0,E MTAB 9860:R968,0:50.48 1:0,A:445006547;C:421206398;G:404638732;T:461295343;N:2502717,50,0,,,445006547,421206398,404638732,461295343,2502717,ERX4788148,ERS5455113,ERA3190565,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93808,,0.10133,,0.65206,,0.47081,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10136,ERR4968950,ERX4788147,ERS5455112,ERP125627,PRJEB41792,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E-MTAB-9860,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of synthetic organochloride insecticide Methoxychlor CAS 72 43 5. It is structurally highly similar to its precursor molecule DDT Dichlorodiphenyltrichloroethane. Like its precursor today Methoxychlor is banned from the use as pesticide in the United States and the European Union due to its acute toxicity high bioaccumulation potential and endocrine disruption activity. The Insecticide Resistance Action Committee IRAC classified Methoxychlor post its mode of action MoA in the target organism as a sodium channel modulator Group 3B. Our goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms for this substance of particularly high environmental concern. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Methoxychlor for xxx hours under semi static conditions. Each test comprised of a low exposure LE 20 µg/L high exposure HE 180 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R964,SAMEA7698597,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698597|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:31Z|INSDC last update:2021 10 15T00:15:31Z|INSDC status:public|Submitter Id:E MTAB 9860:R964|age:96|broker name:ArrayExpress|cas:72 43 5|common name:zebrafish|condition:LowExposure|developmental stage:larval day 4|extraction date:17.02.2020|genotype:wild type genotype|nominal conc:0 00002|organism part:whole larvae|pooled individuals:10|rin:9 1|sample name:E MTAB 9860:R964|sex:male & female pooled|strain:AB|tank spawning group:T10,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E MTAB 9860:R964 s,R964 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:LowExposure|Experimental Factor: dose:0 00002|Experimental Factor: nominal conc:0 00002|Experimental Factor: compound:methoxychlor,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125627,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p915sR964_sr.fastq.gz,fastq,1501519140.0,29746691.0,E MTAB 9860:R964,0:50.48 1:0,A:384276865;C:366094953;G:350798336;T:398037461;N:2311525,50,0,,,384276865,366094953,350798336,398037461,2311525,ERX4788147,ERS5455112,ERA3190565,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93819,,0.09886,,0.65202,,0.46428,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10137,ERR4968949,ERX4788146,ERS5455111,ERP125627,PRJEB41792,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E-MTAB-9860,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of synthetic organochloride insecticide Methoxychlor CAS 72 43 5. It is structurally highly similar to its precursor molecule DDT Dichlorodiphenyltrichloroethane. Like its precursor today Methoxychlor is banned from the use as pesticide in the United States and the European Union due to its acute toxicity high bioaccumulation potential and endocrine disruption activity. The Insecticide Resistance Action Committee IRAC classified Methoxychlor post its mode of action MoA in the target organism as a sodium channel modulator Group 3B. Our goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms for this substance of particularly high environmental concern. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Methoxychlor for xxx hours under semi static conditions. Each test comprised of a low exposure LE 20 µg/L high exposure HE 180 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R960,SAMEA7698596,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698596|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:31Z|INSDC last update:2021 10 15T00:15:31Z|INSDC status:public|Submitter Id:E MTAB 9860:R960|age:96|broker name:ArrayExpress|cas:72 43 5|common name:zebrafish|condition:LowExposure|developmental stage:larval day 4|extraction date:17.02.2020|genotype:wild type genotype|nominal conc:0 00002|organism part:whole larvae|pooled individuals:10|rin:10|sample name:E MTAB 9860:R960|sex:male & female pooled|strain:AB|tank spawning group:T7,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E MTAB 9860:R960 s,R960 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:LowExposure|Experimental Factor: dose:0 00002|Experimental Factor: nominal conc:0 00002|Experimental Factor: compound:methoxychlor,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125627,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p915sR960_sr.fastq.gz,fastq,1729955621.0,34271158.0,E MTAB 9860:R960,0:50.48 1:0,A:440768821;C:423049045;G:407404143;T:456090103;N:2643509,50,0,,,440768821,423049045,407404143,456090103,2643509,ERX4788146,ERS5455111,ERA3190565,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93855,,0.09224,,0.6523,,0.48216,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10138,ERR4968948,ERX4788145,ERS5455110,ERP125627,PRJEB41792,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E-MTAB-9860,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of synthetic organochloride insecticide Methoxychlor CAS 72 43 5. It is structurally highly similar to its precursor molecule DDT Dichlorodiphenyltrichloroethane. Like its precursor today Methoxychlor is banned from the use as pesticide in the United States and the European Union due to its acute toxicity high bioaccumulation potential and endocrine disruption activity. The Insecticide Resistance Action Committee IRAC classified Methoxychlor post its mode of action MoA in the target organism as a sodium channel modulator Group 3B. Our goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms for this substance of particularly high environmental concern. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Methoxychlor for xxx hours under semi static conditions. Each test comprised of a low exposure LE 20 µg/L high exposure HE 180 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R967,SAMEA7698595,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698595|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:31Z|INSDC last update:2021 10 15T00:15:31Z|INSDC status:public|Submitter Id:E MTAB 9860:R967|age:96|broker name:ArrayExpress|cas:72 43 5|common name:zebrafish|condition:Control|developmental stage:larval day 4|extraction date:17.02.2020|genotype:wild type genotype|nominal conc:0|organism part:whole larvae|pooled individuals:10|rin:8 6|sample name:E MTAB 9860:R967|sex:male & female pooled|strain:AB|tank spawning group:T11,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E MTAB 9860:R967 s,R967 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:Control|Experimental Factor: dose:0|Experimental Factor: nominal conc:0|Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125627,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p915sR967_sr.fastq.gz,fastq,1703480505.0,33748366.0,E MTAB 9860:R967,0:50.48 1:0,A:434721421;C:415774473;G:400487304;T:449834359;N:2662948,50,0,,,434721421,415774473,400487304,449834359,2662948,ERX4788145,ERS5455110,ERA3190565,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.9397,,0.09627,,0.6533,,0.47444,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10139,ERR4968947,ERX4788144,ERS5455109,ERP125627,PRJEB41792,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E-MTAB-9860,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of synthetic organochloride insecticide Methoxychlor CAS 72 43 5. It is structurally highly similar to its precursor molecule DDT Dichlorodiphenyltrichloroethane. Like its precursor today Methoxychlor is banned from the use as pesticide in the United States and the European Union due to its acute toxicity high bioaccumulation potential and endocrine disruption activity. The Insecticide Resistance Action Committee IRAC classified Methoxychlor post its mode of action MoA in the target organism as a sodium channel modulator Group 3B. Our goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms for this substance of particularly high environmental concern. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Methoxychlor for xxx hours under semi static conditions. Each test comprised of a low exposure LE 20 µg/L high exposure HE 180 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R963,SAMEA7698594,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698594|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:31Z|INSDC last update:2021 10 15T00:15:31Z|INSDC status:public|Submitter Id:E MTAB 9860:R963|age:96|broker name:ArrayExpress|cas:72 43 5|common name:zebrafish|condition:Control|developmental stage:larval day 4|extraction date:17.02.2020|genotype:wild type genotype|nominal conc:0|organism part:whole larvae|pooled individuals:10|rin:9 4|sample name:E MTAB 9860:R963|sex:male & female pooled|strain:AB|tank spawning group:T10,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E MTAB 9860:R963 s,R963 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:Control|Experimental Factor: dose:0|Experimental Factor: nominal conc:0|Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125627,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p915sR963_sr.fastq.gz,fastq,2049141213.0,40591661.0,E MTAB 9860:R963,0:50.48 1:0,A:523387891;C:498749728;G:482748418;T:541454707;N:2800469,50,0,,,523387891,498749728,482748418,541454707,2800469,ERX4788144,ERS5455109,ERA3190565,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93818,,0.09447,,0.65299,,0.48779,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10140,ERR4968946,ERX4788143,ERS5455108,ERP125627,PRJEB41792,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E-MTAB-9860,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sub lethal concentrations of synthetic organochloride insecticide Methoxychlor CAS 72 43 5. It is structurally highly similar to its precursor molecule DDT Dichlorodiphenyltrichloroethane. Like its precursor today Methoxychlor is banned from the use as pesticide in the United States and the European Union due to its acute toxicity high bioaccumulation potential and endocrine disruption activity. The Insecticide Resistance Action Committee IRAC classified Methoxychlor post its mode of action MoA in the target organism as a sodium channel modulator Group 3B. Our goal is to identify toxicogenomic profiles with predictive character and potential molecular key events KE explaining upstream adverse effects in aquatic non target organisms for this substance of particularly high environmental concern. This will provide useful information to refine and improve existing adverse outcome pathways AOP. Furthermore integrating the obtained profiles for this and other tested chemicals in a collective database will enable us in the future to derive predictions about the ecotoxicological hazard for chemcials with unknown apical effects based on similarly altered transcriptomic and proteomic profiles. In a modified version of the zebrafish embryo toxicity test OECD 236 15 fertilized eggs were exposed to two different sub lethal concentrations of Methoxychlor for xxx hours under semi static conditions. Each test comprised of a low exposure LE 20 µg/L high exposure HE 180 µg/L and negative control NC group and was performed in triplicates. At 96 hpf 10 larvae were randomly picked for each sample and pooled for RNA and protein extraction with NucleoSpin RNA/Protein kit Macherey Nagel. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina HiSeq 4000 System Illumina in 50 bp single read mode producing roughly 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the D.rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R959,SAMEA7698593,"Fraunhofer Attract Eco'n'OMICs, Fraunhofer Institute for Molecular Biology and Applied Ecology, Schmallenberg, Germany Evolutionary Ecology and Environmental Toxicology, Faculty Biological Sciences, Goethe University Frankfurt, Frankfurt, Germany",ENA first public:2021 10 15|ENA last update:2021 10 15|External Id:SAMEA7698593|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology Schmallenberg Germany Evolutionary Ecology and Environmental Toxicology Faculty Biological Sciences Goethe University Frankfurt Frankfurt Germany|INSDC first public:2021 10 15T00:15:31Z|INSDC last update:2021 10 15T00:15:31Z|INSDC status:public|Submitter Id:E MTAB 9860:R959|age:96|broker name:ArrayExpress|cas:72 43 5|common name:zebrafish|condition:Control|developmental stage:larval day 4|extraction date:17.02.2020|genotype:wild type genotype|nominal conc:0|organism part:whole larvae|pooled individuals:10|rin:9 5|sample name:E MTAB 9860:R959|sex:male & female pooled|strain:AB|tank spawning group:T7,,,,,,,,,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,E MTAB 9860:R959 s,R959 s,mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. Methoxychlor CAS 72 43 5 PESTANAL® was purchased from Merck KGgA Darmstadt Germany. For the test solution preparation first a high concentrated stock solution was prepared in Acetone 99.9% MS grade. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle and filled up with pure Acetone to a final volume of 1 ml. To the bottle for the control solution only 1ml of pure Acetone was added. The solvent was then evaporated at room temperature until no Acetone was left in the bottles. At last 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: condition:Control|Experimental Factor: dose:0|Experimental Factor: nominal conc:0|Experimental Factor: compound:n1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP125627,Illumina HiSeq 4000 sequencing; mRNA Seq of zebrafish embryos 96hpf exposed to different concentrations of the organochloride Methoxychlor below acute toxicity levels against untreated control groups,ENA FIRST PUBLIC:2021 10 15|ENA LAST UPDATE:2021 10 15,p915sR959_sr.fastq.gz,fastq,1705014309.0,33812466.0,E MTAB 9860:R959,0:50.43 1:0,A:432620032;C:414853500;G:401540460;T:449275103;N:6725214,50,0,,,432620032,414853500,401540460,449275103,6725214,ERX4788143,ERS5455108,ERA3190565,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93562,,0.09093,,0.65313,,0.47239,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-15,Larval,Larval,Whole Organism,All anatomical structures 10222,ERR6690618,ERX6315387,ERS7605283,ERP131755,PRJEB47482,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD,E-MTAB-10922,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sublethal concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD. MDTETD and THADD both are degradation products of bile salts which are steroid compounds from the digestive tracts of vertebrates which enter the environment upon excretion e.g. in manure.,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1335,SAMEA9926936,Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME,ENA first public:2021 10 11|ENA last update:2021 10 11|External Id:SAMEA9926936|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC first public:2021 10 11T16:25:38Z|INSDC last update:2021 10 11T16:25:38Z|INSDC status:public|Submitter Id:E MTAB 10922:R1335|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|individual:10|organism part:whole organism|sample name:E MTAB 10922:R1335|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to 2 subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1 MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 di1 THADD,E MTAB 10922:R1335 s,R1335 s,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:12β trihydroxy androsta 4 6 diene 3 17 di1|Experimental Factor: dose:1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP131755,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,R1335.fastq.gz,fastq,1688675473.0,33437119.0,E MTAB 10922:R1335,0:50.50 1:0,A:432146956;C:413222132;G:392920682;T:448830781;N:1554922,50,0,,,432146956,413222132,392920682,448830781,1554922,ERX6315387,ERS7605283,ERA6149792,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93124,,0.10466,,0.65249,,0.47759,,48,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-11,Larval,Larval,Whole Organism,All anatomical structures 10223,ERR6690617,ERX6315386,ERS7605282,ERP131755,PRJEB47482,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD,E-MTAB-10922,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sublethal concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD. MDTETD and THADD both are degradation products of bile salts which are steroid compounds from the digestive tracts of vertebrates which enter the environment upon excretion e.g. in manure.,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1331,SAMEA9926935,Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME,ENA first public:2021 10 11|ENA last update:2021 10 11|External Id:SAMEA9926935|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC first public:2021 10 11T16:25:38Z|INSDC last update:2021 10 11T16:25:38Z|INSDC status:public|Submitter Id:E MTAB 10922:R1331|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|individual:10|organism part:whole organism|sample name:E MTAB 10922:R1331|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to 2 subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1 MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 di1 THADD,E MTAB 10922:R1331 s,R1331 s,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:12β trihydroxy androsta 4 6 diene 3 17 di1|Experimental Factor: dose:1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP131755,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,R1331.fastq.gz,fastq,2114660599.0,41862178.0,E MTAB 10922:R1331,0:50.51 1:0,A:538714643;C:520186712;G:495375016;T:559484406;N:899822,50,0,,,538714643,520186712,495375016,559484406,899822,ERX6315386,ERS7605282,ERA6149792,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.92976,,0.09702,,0.65419,,0.45924,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-11,Larval,Larval,Whole Organism,All anatomical structures 10224,ERR6690616,ERX6315385,ERS7605281,ERP131755,PRJEB47482,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD,E-MTAB-10922,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sublethal concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD. MDTETD and THADD both are degradation products of bile salts which are steroid compounds from the digestive tracts of vertebrates which enter the environment upon excretion e.g. in manure.,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1327,SAMEA9926934,Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME,ENA first public:2021 10 11|ENA last update:2021 10 11|External Id:SAMEA9926934|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC first public:2021 10 11T16:25:38Z|INSDC last update:2021 10 11T16:25:38Z|INSDC status:public|Submitter Id:E MTAB 10922:R1327|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|individual:10|organism part:whole organism|sample name:E MTAB 10922:R1327|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to 2 subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1 MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 di1 THADD,E MTAB 10922:R1327 s,R1327 s,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:12β trihydroxy androsta 4 6 diene 3 17 di1|Experimental Factor: dose:1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP131755,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,R1327.fastq.gz,fastq,1779358537.0,35230725.0,E MTAB 10922:R1327,0:50.51 1:0,A:455709630;C:435432679;G:414075559;T:472716969;N:1423700,50,0,,,455709630,435432679,414075559,472716969,1423700,ERX6315385,ERS7605281,ERA6149792,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.92609,,0.10344,,0.65052,,0.47367,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-11,Larval,Larval,Whole Organism,All anatomical structures 10225,ERR6690615,ERX6315384,ERS7605280,ERP131755,PRJEB47482,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD,E-MTAB-10922,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sublethal concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD. MDTETD and THADD both are degradation products of bile salts which are steroid compounds from the digestive tracts of vertebrates which enter the environment upon excretion e.g. in manure.,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1334,SAMEA9926933,Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME,ENA first public:2021 10 11|ENA last update:2021 10 11|External Id:SAMEA9926933|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC first public:2021 10 11T16:25:38Z|INSDC last update:2021 10 11T16:25:38Z|INSDC status:public|Submitter Id:E MTAB 10922:R1334|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|individual:10|organism part:whole organism|sample name:E MTAB 10922:R1334|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to 2 subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1 MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 di1 THADD,E MTAB 10922:R1334 s,R1334 s,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:12β trihydroxy androsta 4 6 diene 3 17 di1|Experimental Factor: dose:0.1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP131755,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,R1334.fastq.gz,fastq,1808121386.0,35796800.0,E MTAB 10922:R1334,0:50.51 1:0,A:460840812;C:444321139;G:423234972;T:478828245;N:896218,50,0,,,460840812,444321139,423234972,478828245,896218,ERX6315384,ERS7605280,ERA6149792,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.92927,,0.09981,,0.6534,,0.47641,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-11,Larval,Larval,Whole Organism,All anatomical structures 10226,ERR6690614,ERX6315383,ERS7605279,ERP131755,PRJEB47482,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD,E-MTAB-10922,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sublethal concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD. MDTETD and THADD both are degradation products of bile salts which are steroid compounds from the digestive tracts of vertebrates which enter the environment upon excretion e.g. in manure.,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1330,SAMEA9926932,Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME,ENA first public:2021 10 11|ENA last update:2021 10 11|External Id:SAMEA9926932|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC first public:2021 10 11T16:25:38Z|INSDC last update:2021 10 11T16:25:38Z|INSDC status:public|Submitter Id:E MTAB 10922:R1330|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|individual:10|organism part:whole organism|sample name:E MTAB 10922:R1330|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to 2 subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1 MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 di1 THADD,E MTAB 10922:R1330 s,R1330 s,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:12β trihydroxy androsta 4 6 diene 3 17 di1|Experimental Factor: dose:0.1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP131755,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,R1330.fastq.gz,fastq,1493810880.0,29573271.0,E MTAB 10922:R1330,0:50.51 1:0,A:382619085;C:365079492;G:347711071;T:397843564;N:557668,50,0,,,382619085,365079492,347711071,397843564,557668,ERX6315383,ERS7605279,ERA6149792,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.92929,,0.09909,,0.65054,,0.47332,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-11,Larval,Larval,Whole Organism,All anatomical structures 10227,ERR6690613,ERX6315382,ERS7605278,ERP131755,PRJEB47482,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD,E-MTAB-10922,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sublethal concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD. MDTETD and THADD both are degradation products of bile salts which are steroid compounds from the digestive tracts of vertebrates which enter the environment upon excretion e.g. in manure.,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1326,SAMEA9926931,Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME,ENA first public:2021 10 11|ENA last update:2021 10 11|External Id:SAMEA9926931|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC first public:2021 10 11T16:25:38Z|INSDC last update:2021 10 11T16:25:38Z|INSDC status:public|Submitter Id:E MTAB 10922:R1326|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|individual:10|organism part:whole organism|sample name:E MTAB 10922:R1326|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to 2 subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1 MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 di1 THADD,E MTAB 10922:R1326 s,R1326 s,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:12β trihydroxy androsta 4 6 diene 3 17 di1|Experimental Factor: dose:0.1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP131755,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,R1326.fastq.gz,fastq,1556362878.0,30814409.0,E MTAB 10922:R1326,0:50.51 1:0,A:399138616;C:379536887;G:362721177;T:413983572;N:982626,50,0,,,399138616,379536887,362721177,413983572,982626,ERX6315382,ERS7605278,ERA6149792,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.92334,,0.10268,,0.65206,,0.47006,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-11,Larval,Larval,Whole Organism,All anatomical structures 10228,ERR6690612,ERX6315381,ERS7605277,ERP131755,PRJEB47482,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD,E-MTAB-10922,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sublethal concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD. MDTETD and THADD both are degradation products of bile salts which are steroid compounds from the digestive tracts of vertebrates which enter the environment upon excretion e.g. in manure.,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1332,SAMEA9926930,Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME,ENA first public:2021 10 11|ENA last update:2021 10 11|External Id:SAMEA9926930|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC first public:2021 10 11T16:25:38Z|INSDC last update:2021 10 11T16:25:38Z|INSDC status:public|Submitter Id:E MTAB 10922:R1332|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|individual:10|organism part:whole organism|sample name:E MTAB 10922:R1332|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to 2 subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1 MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 di1 THADD,E MTAB 10922:R1332 s,R1332 s,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:n1|Experimental Factor: dose:0,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP131755,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,R1332.fastq.gz,fastq,1724677047.0,34142889.0,E MTAB 10922:R1332,0:50.51 1:0,A:441607887;C:421814110;G:401384804;T:459230039;N:640207,50,0,,,441607887,421814110,401384804,459230039,640207,ERX6315381,ERS7605277,ERA6149792,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.92138,,0.10053,,0.65399,,0.47486,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-11,Larval,Larval,Whole Organism,All anatomical structures 10229,ERR6690611,ERX6315380,ERS7605276,ERP131755,PRJEB47482,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD,E-MTAB-10922,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sublethal concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD. MDTETD and THADD both are degradation products of bile salts which are steroid compounds from the digestive tracts of vertebrates which enter the environment upon excretion e.g. in manure.,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1328,SAMEA9926929,Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME,ENA first public:2021 10 11|ENA last update:2021 10 11|External Id:SAMEA9926929|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC first public:2021 10 11T16:25:38Z|INSDC last update:2021 10 11T16:25:38Z|INSDC status:public|Submitter Id:E MTAB 10922:R1328|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|individual:10|organism part:whole organism|sample name:E MTAB 10922:R1328|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to 2 subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1 MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 di1 THADD,E MTAB 10922:R1328 s,R1328 s,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:n1|Experimental Factor: dose:0,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP131755,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,R1328.fastq.gz,fastq,2338196773.0,46290960.0,E MTAB 10922:R1328,0:50.51 1:0,A:597574623;C:571603200;G:546389543;T:621576532;N:1052875,50,0,,,597574623,571603200,546389543,621576532,1052875,ERX6315380,ERS7605276,ERA6149792,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.9274,,0.10154,,0.65139,,0.47399,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-11,Larval,Larval,Whole Organism,All anatomical structures 10230,ERR6690610,ERX6315379,ERS7605275,ERP131755,PRJEB47482,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD,E-MTAB-10922,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sublethal concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD. MDTETD and THADD both are degradation products of bile salts which are steroid compounds from the digestive tracts of vertebrates which enter the environment upon excretion e.g. in manure.,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1324,SAMEA9926928,Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME,ENA first public:2021 10 11|ENA last update:2021 10 11|External Id:SAMEA9926928|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC first public:2021 10 11T16:25:38Z|INSDC last update:2021 10 11T16:25:38Z|INSDC status:public|Submitter Id:E MTAB 10922:R1324|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|individual:10|organism part:whole organism|sample name:E MTAB 10922:R1324|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to 2 subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1 MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 di1 THADD,E MTAB 10922:R1324 s,R1324 s,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:n1|Experimental Factor: dose:0,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP131755,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,R1324.fastq.gz,fastq,1661447208.0,32893507.0,E MTAB 10922:R1324,0:50.51 1:0,A:426380918;C:404798726;G:385491952;T:444113172;N:662440,50,0,,,426380918,404798726,385491952,444113172,662440,ERX6315379,ERS7605275,ERA6149792,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93256,,0.09976,,0.64597,,0.47142,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-11,Larval,Larval,Whole Organism,All anatomical structures 10231,ERR6690609,ERX6315378,ERS7605274,ERP131755,PRJEB47482,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD,E-MTAB-10922,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sublethal concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD. MDTETD and THADD both are degradation products of bile salts which are steroid compounds from the digestive tracts of vertebrates which enter the environment upon excretion e.g. in manure.,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1347,SAMEA9926927,Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME,ENA first public:2021 10 11|ENA last update:2021 10 11|External Id:SAMEA9926927|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC first public:2021 10 11T16:25:38Z|INSDC last update:2021 10 11T16:25:38Z|INSDC status:public|Submitter Id:E MTAB 10922:R1347|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|individual:10|organism part:whole organism|sample name:E MTAB 10922:R1347|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to 2 subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1 MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 di1 THADD,E MTAB 10922:R1347 s,R1347 s,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1|Experimental Factor: dose:1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP131755,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,R1347.fastq.gz,fastq,2013684171.0,39865705.0,E MTAB 10922:R1347,0:50.51 1:0,A:513405372;C:495540575;G:469675681;T:534136644;N:925899,50,0,,,513405372,495540575,469675681,534136644,925899,ERX6315378,ERS7605274,ERA6149792,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.93259,,0.09724,,0.65269,,0.46848,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-11,Larval,Larval,Whole Organism,All anatomical structures 10232,ERR6690608,ERX6315377,ERS7605273,ERP131755,PRJEB47482,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD,E-MTAB-10922,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sublethal concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD. MDTETD and THADD both are degradation products of bile salts which are steroid compounds from the digestive tracts of vertebrates which enter the environment upon excretion e.g. in manure.,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1343,SAMEA9926926,Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME,ENA first public:2021 10 11|ENA last update:2021 10 11|External Id:SAMEA9926926|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC first public:2021 10 11T16:25:38Z|INSDC last update:2021 10 11T16:25:38Z|INSDC status:public|Submitter Id:E MTAB 10922:R1343|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|individual:10|organism part:whole organism|sample name:E MTAB 10922:R1343|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to 2 subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1 MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 di1 THADD,E MTAB 10922:R1343 s,R1343 s,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1|Experimental Factor: dose:1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP131755,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,R1343.fastq.gz,fastq,2126807115.0,42106048.0,E MTAB 10922:R1343,0:50.51 1:0,A:547291227;C:518441995;G:491529136;T:568496689;N:1048068,50,0,,,547291227,518441995,491529136,568496689,1048068,ERX6315377,ERS7605273,ERA6149792,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.92904,,0.10754,,0.64999,,0.47787,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-11,Larval,Larval,Whole Organism,All anatomical structures 10233,ERR6690607,ERX6315376,ERS7605272,ERP131755,PRJEB47482,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD,E-MTAB-10922,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sublethal concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD. MDTETD and THADD both are degradation products of bile salts which are steroid compounds from the digestive tracts of vertebrates which enter the environment upon excretion e.g. in manure.,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1339,SAMEA9926925,Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME,ENA first public:2021 10 11|ENA last update:2021 10 11|External Id:SAMEA9926925|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC first public:2021 10 11T16:25:38Z|INSDC last update:2021 10 11T16:25:38Z|INSDC status:public|Submitter Id:E MTAB 10922:R1339|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|individual:10|organism part:whole organism|sample name:E MTAB 10922:R1339|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to 2 subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1 MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 di1 THADD,E MTAB 10922:R1339 s,R1339 s,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1|Experimental Factor: dose:1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP131755,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,R1339.fastq.gz,fastq,1877735463.0,37173221.0,E MTAB 10922:R1339,0:50.51 1:0,A:485012954;C:455923798;G:432567831;T:503496745;N:734135,50,0,,,485012954,455923798,432567831,503496745,734135,ERX6315376,ERS7605272,ERA6149792,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.92237,,0.11315,,0.65313,,0.46728,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-11,Larval,Larval,Whole Organism,All anatomical structures 10234,ERR6690606,ERX6315375,ERS7605271,ERP131755,PRJEB47482,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD,E-MTAB-10922,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sublethal concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD. MDTETD and THADD both are degradation products of bile salts which are steroid compounds from the digestive tracts of vertebrates which enter the environment upon excretion e.g. in manure.,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1346,SAMEA9926924,Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME,ENA first public:2021 10 11|ENA last update:2021 10 11|External Id:SAMEA9926924|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC first public:2021 10 11T16:25:38Z|INSDC last update:2021 10 11T16:25:38Z|INSDC status:public|Submitter Id:E MTAB 10922:R1346|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|individual:10|organism part:whole organism|sample name:E MTAB 10922:R1346|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to 2 subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1 MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 di1 THADD,E MTAB 10922:R1346 s,R1346 s,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1|Experimental Factor: dose:0.1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP131755,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,R1346.fastq.gz,fastq,2055618983.0,40694887.0,E MTAB 10922:R1346,0:50.51 1:0,A:526660468;C:501970997;G:479146997;T:546796841;N:1043680,50,0,,,526660468,501970997,479146997,546796841,1043680,ERX6315375,ERS7605271,ERA6149792,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.92338,,0.10341,,0.65541,,0.47789,,50,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-11,Larval,Larval,Whole Organism,All anatomical structures 10235,ERR6690605,ERX6315374,ERS7605270,ERP131755,PRJEB47482,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD,E-MTAB-10922,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sublethal concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD. MDTETD and THADD both are degradation products of bile salts which are steroid compounds from the digestive tracts of vertebrates which enter the environment upon excretion e.g. in manure.,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1342,SAMEA9926923,Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME,ENA first public:2021 10 11|ENA last update:2021 10 11|External Id:SAMEA9926923|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC first public:2021 10 11T16:25:38Z|INSDC last update:2021 10 11T16:25:38Z|INSDC status:public|Submitter Id:E MTAB 10922:R1342|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|individual:10|organism part:whole organism|sample name:E MTAB 10922:R1342|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to 2 subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1 MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 di1 THADD,E MTAB 10922:R1342 s,R1342 s,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1|Experimental Factor: dose:0.1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP131755,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,R1342.fastq.gz,fastq,1805917169.0,35751110.0,E MTAB 10922:R1342,0:50.51 1:0,A:464432790;C:439427848;G:418569252;T:482744908;N:742371,50,0,,,464432790,439427848,418569252,482744908,742371,ERX6315374,ERS7605270,ERA6149792,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.92785,,0.10931,,0.65265,,0.46925,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-11,Larval,Larval,Whole Organism,All anatomical structures 10236,ERR6690604,ERX6315373,ERS7605269,ERP131755,PRJEB47482,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD,E-MTAB-10922,Transcriptome Analysis,The aim of this mRNA expression profiling experiment was to screen for ecotoxicogenomic fingerprints in zebrafish Danio rerio embryos as aquatic vertebrate non target model exposed to sublethal concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12ß trihydroxy androsta 4 6 diene 3 17 dione THADD. MDTETD and THADD both are degradation products of bile salts which are steroid compounds from the digestive tracts of vertebrates which enter the environment upon excretion e.g. in manure.,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,,Protocols: For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,R1338,SAMEA9926922,Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME,ENA first public:2021 10 11|ENA last update:2021 10 11|External Id:SAMEA9926922|INSDC center alias:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC center name:Fraunhofer Attract Eco'n'OMICs Fraunhofer Institute for Molecular Biology and Applied Ecology IME|INSDC first public:2021 10 11T16:25:38Z|INSDC last update:2021 10 11T16:25:38Z|INSDC status:public|Submitter Id:E MTAB 10922:R1338|age:96|broker name:ArrayExpress|common name:zebrafish|developmental stage:larval day 4|genotype:wild type genotype|individual:10|organism part:whole organism|sample name:E MTAB 10922:R1338|sex:mixed|strain:AB,,,,,,,,,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to 2 subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1 MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 di1 THADD,E MTAB 10922:R1338 s,R1338 s,Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,For each sample 10 dpf 4 dpf zebrafish larvae were randomly picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Once individuals were collected for all samples they were euthanized at the same time by placing the tubes simultaneously on ice for 10 minutes. Supernatant from the test solutions was then carefully removed without xxx the larvae before adding 350µl of RP1 buffer NucleoSpin RNA Protein kit; Macherey Nagel 740933.250 for tissue homogenization. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: Wild type zebrafish Danio rerio Strain AB broodstocks were maintained under flow through conditions in 150 L tanks at 26 +/ 2°C on a 12:12 h light/dark cycle. They were fed daily with TetraMin® Tetra Werke Melle Germany main feed ad libitum and nauplii of Artemia salina. The day before test start glass spawning trays with artificial substrate green glass beads stringed on stainless steel wire were placed at the bottom of each tank. post mating and spawning in the morning hours eggs were rinsed with clear Cu reduced water and placed into glass dishes for pre picking. Fish embryo incubation: For each sample 15 fertilized eggs in the early blastula stage were placed in glass petri dishes diameter 6 cm filled with 8 ml of the respective testing solution Control Low exposure High exposure. Embryos were incubated at 27 +/ 1°C on a 14:10 h light/dark cycle. At 24 hpf eggs were inspected visually and single coagulated eggs were recorded and removed. At 48 hpf overall survival hatch rates morphological malformations and physiological changes were recorded and aged solutions were replaced by fresh aerated test solutions. At 96 hpf again overall survival hatch rates morphological malformations and physiological changes were recorded before RNA and protein extraction. For the test solution preparation first a high concentrated stock solution was prepared in water. From this stock respective volumes for each testing concentration were added to a 250 ml DURAN glass bottle. 200 ml of Cu reduced water were added and solutions were stirred in the dark for 3 hours at room temperature. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 96 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. Per sample about 40 50 fertilized fish eggs were pre picked in glass petri dishes 10 cm diameter filled with 30 ml of the testing solution. Using an optical binokular microscope 15 embryos of the same early blastula stage were transferred from the pre picking plate into the test glass wells filled with 8 ml of the test solution. Embryos were then incubated as described in the “growth protocol”. The pooled larvae in RP1 buffer were transferred to a screw cap eppendorf filled with 0.3 g of Lysing Matrix D MP Biomedicals 6913050 ceramic beads. Tissue homogenization was performed at 5 m/s for 45 s with FastPrep 24© MP Biomedicals 6004500 at room temperature. From here on total RNA and Protein was extracted from the tissue lysate using the NucleoSpin RNA Protein kit Macherey Nagel 740933.250 according to the manufacturer's protocol. RNA concentration >100 ng/µl was measured using a Nanodrop 2000 spectrophotometer Thermo Scientific. The sample's overall RNA quality and corresponding RIN value was fluorometrically determined with a 2100 Bioanalyzer© Instrument system Agilent G2939BA using the Agilent RNA 6000 Nano Kit© Agilent 5067 1511 according to the manufacturer's protocol. Only samples with a RIN > 8 were used for downstream analysis. Samples were stored at 80°C until they were send to the sequencing facility on dry ice. Sequencing libraries were prepared for each sample from 100 ng/µl total RNA at the sequencing facility “NGS Services for Integrative Genomics” at the University of Göttingen in Germany. According to their standard workflow cDNA libraries were prepared from protein coding mRNA that were purified through PolyA selection using the TruSeq RNA Library Prep Kit v2 Illumina. Libraries were validated using a Fragment Analyzer system Agilent Santa Clara USA before sequencing.,Experimental Factor: compound:4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 di1|Experimental Factor: dose:0.1,RNA-Seq,TRANSCRIPTOMIC,PolyA,SINGLE,ILLUMINA,Illumina HiSeq 4000,,ERP131755,Illumina HiSeq 4000 sequencing; Transcriptional profiling of zebrafish embryos post 96 h exposure to two subsequent concentrations of 4 methyl 3 deoxy 1 9 12 trihydroxyestra 1 3 5107 tetraene 6 17 dione MDTETD and 12β trihydroxy androsta 4 6 diene 3 17 dione THADD,ENA FIRST PUBLIC:2021 10 11|ENA LAST UPDATE:2021 10 11,R1338.fastq.gz,fastq,863193301.0,17089189.0,E MTAB 10922:R1338,0:50.51 1:0,A:221094712;C:211145297;G:201026970;T:229486432;N:439890,50,0,,,221094712,211145297,201026970,229486432,439890,ERX6315373,ERS7605269,ERA6149792,Fraunhofer Attract Eco,Fraunhofer Attract Eco,1,0.92768,,0.103,,0.65145,,0.47316,,51,,B,,usable mapping rate,illumina,hiseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,Germany,2021-10-11,Larval,Larval,Whole Organism,All anatomical structures