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
60,DRR032764,DRX029570,DRS049969,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 100 individuals,Dr shield 2,SAMD00028161,,sample name:Dr shield 2|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:shield|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028161,DRX029570,Dr shield 2,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028161,,,,3644397900.0,36443979.0,DRR032764,0:100 1:0,A:986071173;C:842367218;G:837686080;T:978236607;N:36822,100,0,,,986071173,842367218,837686080,978236607,36822,DRX029570,DRS049969,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92419,,0.08269,,0.75558,,0.47863,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Gastrula,Embryo,Whole Organism,All anatomical structures
61,DRR032763,DRX029569,DRS049968,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 100 individuals,Dr shield 1,SAMD00028160,,sample name:Dr shield 1|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:shield|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028160,DRX029569,Dr shield 1,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028160,,,,3834622000.0,38346220.0,DRR032763,0:100 1:0,A:1043352851;C:880011834;G:876775415;T:1034444253;N:37647,100,0,,,1043352851,880011834,876775415,1034444253,37647,DRX029569,DRS049968,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92305,,0.09126,,0.75481,,0.47587,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Gastrula,Embryo,Whole Organism,All anatomical structures
62,DRR032762,DRX029568,DRS049967,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 50 individuals,Dr prime5 6 3,SAMD00028159,,sample name:Dr prime5 6 3|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:prime5 6|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028159,DRX029568,Dr prime5 6 3,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028159,,,,3903332800.0,39033328.0,DRR032762,0:100 1:0,A:1050045822;C:908538410;G:900588661;T:1044116537;N:43370,100,0,,,1050045822,908538410,900588661,1044116537,43370,DRX029568,DRS049967,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92761,,0.07976,,0.69126,,0.46568,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Undetermined,Embryo,Whole Organism,All anatomical structures
63,DRR032761,DRX029567,DRS049966,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 50 individuals,Dr prime5 6 2,SAMD00028158,,sample name:Dr prime5 6 2|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:prime5 6|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028158,DRX029567,Dr prime5 6 2,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028158,,,,3678549700.0,36785497.0,DRR032761,0:100 1:0,A:986526644;C:857762765;G:853417738;T:980801764;N:40789,100,0,,,986526644,857762765,853417738,980801764,40789,DRX029567,DRS049966,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92689,,0.07872,,0.6928,,0.46577,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Undetermined,Embryo,Whole Organism,All anatomical structures
64,DRR032760,DRX029566,DRS049965,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 50 individuals,Dr prime5 6 1,SAMD00028157,,sample name:Dr prime5 6 1|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:prime5 6|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028157,DRX029566,Dr prime5 6 1,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028157,,,,3863129500.0,38631295.0,DRR032760,0:100 1:0,A:1035240477;C:901625010;G:895370149;T:1030851937;N:41927,100,0,,,1035240477,901625010,895370149,1030851937,41927,DRX029566,DRS049965,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92337,,0.07522,,0.69315,,0.46516,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Undetermined,Embryo,Whole Organism,All anatomical structures
65,DRR032759,DRX029565,DRS049964,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 50 individuals,Dr prime25 2,SAMD00028156,,sample name:Dr prime25 2|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:prime25|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028156,DRX029565,Dr prime25 2,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028156,,,,3750136100.0,37501361.0,DRR032759,0:100 1:0,A:1013528040;C:866734984;G:862431819;T:1007403208;N:38049,100,0,,,1013528040,866734984,862431819,1007403208,38049,DRX029565,DRS049964,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92019,,0.09079,,0.68304,,0.47083,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Undetermined,Embryo,Whole Organism,All anatomical structures
66,DRR032758,DRX029564,DRS049963,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 50 individuals,Dr prime25 1,SAMD00028155,,sample name:Dr prime25 1|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:prime25|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028155,DRX029564,Dr prime25 1,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028155,,,,3544862700.0,35448627.0,DRR032758,0:100 1:0,A:952135895;C:825841753;G:821757889;T:945087927;N:39236,100,0,,,952135895,825841753,821757889,945087927,39236,DRX029564,DRS049963,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92229,,0.08344,,0.68525,,0.466,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Undetermined,Embryo,Whole Organism,All anatomical structures
67,DRR032757,DRX029563,DRS049962,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 97 individuals,Dr bud 2,SAMD00028154,,sample name:Dr bud 2|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:bud|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028154,DRX029563,Dr bud 2,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028154,,,,4104778200.0,41047782.0,DRR032757,0:100 1:0,A:1116316188;C:944738800;G:936257056;T:1107423486;N:42670,100,0,,,1116316188,944738800,936257056,1107423486,42670,DRX029563,DRS049962,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92945,,0.10493,,0.73407,,0.47824,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Undetermined,Embryo,Whole Organism,All anatomical structures
68,DRR032756,DRX029562,DRS049961,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 100 individuals,Dr bud 1,SAMD00028153,,sample name:Dr bud 1|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:bud|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028153,DRX029562,Dr bud 1,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028153,,,,4540291000.0,45402910.0,DRR032756,0:100 1:0,A:1237914068;C:1042346110;G:1033172731;T:1226799791;N:58300,100,0,,,1237914068,1042346110,1033172731,1226799791,58300,DRX029562,DRS049961,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92628,,0.10478,,0.7391,,0.46461,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Undetermined,Embryo,Whole Organism,All anatomical structures
69,DRR032755,DRX029561,DRS049960,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 100 individuals,Dr 90epiboly 2,SAMD00028152,,sample name:Dr 90epiboly 2|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:90epiboly|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028152,DRX029561,Dr 90epiboly 2,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028152,,,,3572358600.0,35723586.0,DRR032755,0:100 1:0,A:971653450;C:821326559;G:816855636;T:962477457;N:45498,100,0,,,971653450,821326559,816855636,962477457,45498,DRX029561,DRS049960,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92485,,0.10642,,0.74213,,0.47012,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Gastrula,Embryo,Whole Organism,All anatomical structures
70,DRR032754,DRX029560,DRS049959,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 100 individuals,Dr 90epiboly 1,SAMD00028151,,sample name:Dr 90epiboly 1|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:90epiboly|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028151,DRX029560,Dr 90epiboly 1,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028151,,,,3423980500.0,34239805.0,DRR032754,0:100 1:0,A:933088185;C:785251613;G:780911148;T:924686406;N:43148,100,0,,,933088185,785251613,780911148,924686406,43148,DRX029560,DRS049959,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92436,,0.10881,,0.74255,,0.47068,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Gastrula,Embryo,Whole Organism,All anatomical structures
71,DRR032753,DRX029559,DRS049958,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 114 individuals,Dr 8cell 2,SAMD00028150,,sample name:Dr 8cell 2|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:8cell|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028150,DRX029559,Dr 8cell 2,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028150,,,,3708921900.0,37089219.0,DRR032753,0:100 1:0,A:985502141;C:874161613;G:869551685;T:979663686;N:42775,100,0,,,985502141,874161613,869551685,979663686,42775,DRX029559,DRS049958,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.93329,,0.02366,,0.78896,,0.47447,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Cleavage,Embryo,Whole Organism,All anatomical structures
72,DRR032752,DRX029558,DRS049957,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 96 individuals,Dr 8cell 1,SAMD00028149,,sample name:Dr 8cell 1|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:8cell|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028149,DRX029558,Dr 8cell 1,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028149,,,,3666991200.0,36669912.0,DRR032752,0:100 1:0,A:976118513;C:862559696;G:858017821;T:970254302;N:40868,100,0,,,976118513,862559696,858017821,970254302,40868,DRX029558,DRS049957,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.934,,0.02403,,0.78877,,0.46902,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Cleavage,Embryo,Whole Organism,All anatomical structures
73,DRR032751,DRX029557,DRS049956,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 100 individuals,Dr 75epiboly 2,SAMD00028148,,sample name:Dr 75epiboly 2|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:75epiboly|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028148,DRX029557,Dr 75epiboly 2,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028148,,,,3252021500.0,32520215.0,DRR032751,0:100 1:0,A:885527595;C:746750899;G:742907892;T:876794123;N:40991,100,0,,,885527595,746750899,742907892,876794123,40991,DRX029557,DRS049956,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92594,,0.10181,,0.74862,,0.47789,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Gastrula,Embryo,Whole Organism,All anatomical structures
74,DRR032750,DRX029556,DRS049955,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 100 individuals,Dr 75epiboly 1,SAMD00028147,,sample name:Dr 75epiboly 1|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:75epiboly|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028147,DRX029556,Dr 75epiboly 1,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028147,,,,3785053700.0,37850537.0,DRR032750,0:100 1:0,A:1029014798;C:870946157;G:867537069;T:1017508684;N:46992,100,0,,,1029014798,870946157,867537069,1017508684,46992,DRX029556,DRS049955,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92346,,0.10046,,0.74921,,0.47295,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Gastrula,Embryo,Whole Organism,All anatomical structures
75,DRR032749,DRX029555,DRS049954,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 50 individuals,Dr 72h 2,SAMD00028146,,sample name:Dr 72h 2|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:72h Protruding mouth|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028146,DRX029555,Dr 72h 2,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028146,,,,3429795800.0,34297958.0,DRR032749,0:100 1:0,A:928062015;C:792470305;G:786930881;T:922296289;N:36310,100,0,,,928062015,792470305,786930881,922296289,36310,DRX029555,DRS049954,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.91821,,0.09774,,0.65437,,0.46443,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Larval,Larval,Whole Organism,All anatomical structures
76,DRR032748,DRX029554,DRS049953,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 50 individuals,Dr 72h 1,SAMD00028145,,sample name:Dr 72h 1|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:72h Protruding mouth|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028145,DRX029554,Dr 72h 1,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028145,,,,3897194500.0,38971945.0,DRR032748,0:100 1:0,A:1050989414;C:903225496;G:895783177;T:1047153493;N:42920,100,0,,,1050989414,903225496,895783177,1047153493,42920,DRX029554,DRS049953,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92211,,0.09393,,0.65486,,0.45971,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Larval,Larval,Whole Organism,All anatomical structures
77,DRR032747,DRX029553,DRS049952,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 50 individuals,Dr 6somite 2,SAMD00028144,,sample name:Dr 6somite 2|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:6somite|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028144,DRX029553,Dr 6somite 2,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028144,,,,3704431000.0,37044310.0,DRR032747,0:100 1:0,A:1001844161;C:856702913;G:850695568;T:995148798;N:39560,100,0,,,1001844161,856702913,850695568,995148798,39560,DRX029553,DRS049952,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92633,,0.09211,,0.72107,,0.47195,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Segmentation,Embryo,Whole Organism,All anatomical structures
78,DRR032746,DRX029552,DRS049951,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 50 individuals,Dr 6somite 1,SAMD00028143,,sample name:Dr 6somite 1|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:6somite|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028143,DRX029552,Dr 6somite 1,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028143,,,,3529311900.0,35293119.0,DRR032746,0:100 1:0,A:953957996;C:816824469;G:811403696;T:947089530;N:36209,100,0,,,953957996,816824469,811403696,947089530,36209,DRX029552,DRS049951,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92437,,0.09257,,0.72113,,0.47004,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Segmentation,Embryo,Whole Organism,All anatomical structures
79,DRR032745,DRX029551,DRS049950,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 50 individuals,Dr 60h 2,SAMD00028142,,sample name:Dr 60h 2|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:60h Pec fin|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028142,DRX029551,Dr 60h 2,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028142,,,,3875337000.0,38753370.0,DRR032745,0:100 1:0,A:1042558903;C:899892111;G:896867583;T:1035981420;N:36983,100,0,,,1042558903,899892111,896867583,1035981420,36983,DRX029551,DRS049950,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.91891,,0.09445,,0.66156,,0.45564,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Hatching,Embryo,Whole Organism,All anatomical structures
80,DRR032744,DRX029550,DRS049949,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 50 individuals,Dr 60h 1,SAMD00028141,,sample name:Dr 60h 1|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:60h Pec fin|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028141,DRX029550,Dr 60h 1,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028141,,,,3538468200.0,35384682.0,DRR032744,0:100 1:0,A:960664313;C:812459988;G:809014008;T:956295205;N:34686,100,0,,,960664313,812459988,809014008,956295205,34686,DRX029550,DRS049949,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.91388,,0.10346,,0.66076,,0.45203,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Hatching,Embryo,Whole Organism,All anatomical structures
81,DRR032743,DRX029549,DRS049948,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 50 individuals,Dr 5day 3,SAMD00028140,,sample name:Dr 5day 3|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:5day|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028140,DRX029549,Dr 5day 3,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028140,,,,3884716000.0,38847160.0,DRR032743,0:100 1:0,A:1040550584;C:905663425;G:904247323;T:1034215019;N:39649,100,0,,,1040550584,905663425,904247323,1034215019,39649,DRX029549,DRS049948,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92219,,0.08287,,0.65863,,0.47377,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Larval,Larval,Whole Organism,All anatomical structures
82,DRR032742,DRX029548,DRS049947,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 50 individuals,Dr 5day 2,SAMD00028139,,sample name:Dr 5day 2|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:5day|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028139,DRX029548,Dr 5day 2,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028139,,,,3893708700.0,38937087.0,DRR032742,0:100 1:0,A:1050850168;C:899863467;G:897224776;T:1045729184;N:41105,100,0,,,1050850168,899863467,897224776,1045729184,41105,DRX029548,DRS049947,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.91671,,0.0991,,0.65161,,0.47454,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Larval,Larval,Whole Organism,All anatomical structures
83,DRR032741,DRX029547,DRS049946,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 50 individuals,Dr 5day 1,SAMD00028138,,sample name:Dr 5day 1|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:5day|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028138,DRX029547,Dr 5day 1,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028138,,,,3807570600.0,38075706.0,DRR032741,0:100 1:0,A:1022590228;C:884655401;G:882546091;T:1017737883;N:40997,100,0,,,1022590228,884655401,882546091,1017737883,40997,DRX029547,DRS049946,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.9182,,0.09442,,0.65525,,0.46661,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Larval,Larval,Whole Organism,All anatomical structures
84,DRR032740,DRX029546,DRS049945,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 50 individuals,Dr 48h 2,SAMD00028137,,sample name:Dr 48h 2|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:48h Long pec|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028137,DRX029546,Dr 48h 2,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028137,,,,3702804700.0,37028047.0,DRR032740,0:100 1:0,A:993931475;C:862403562;G:857808891;T:988623734;N:37038,100,0,,,993931475,862403562,857808891,988623734,37038,DRX029546,DRS049945,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92508,,0.08526,,0.68349,,0.45769,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Hatching,Embryo,Whole Organism,All anatomical structures
85,DRR032739,DRX029545,DRS049944,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 50 individuals,Dr 48h 1,SAMD00028136,,sample name:Dr 48h 1|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:48h Long pec|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028136,DRX029545,Dr 48h 1,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028136,,,,3980240400.0,39802404.0,DRR032739,0:100 1:0,A:1070497788;C:925240883;G:920038728;T:1064422474;N:40527,100,0,,,1070497788,925240883,920038728,1064422474,40527,DRX029545,DRS049944,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92349,,0.08681,,0.67874,,0.46565,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Hatching,Embryo,Whole Organism,All anatomical structures
86,DRR032738,DRX029544,DRS049943,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 100 individuals,Dr 32cell 2,SAMD00028135,,sample name:Dr 32cell 2|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:32cell|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028135,DRX029544,Dr 32cell 2,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028135,,,,3678713000.0,36787130.0,DRR032738,0:100 1:0,A:981005900;C:863203049;G:859660640;T:974807835;N:35576,100,0,,,981005900,863203049,859660640,974807835,35576,DRX029544,DRS049943,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.93302,,0.02468,,0.77441,,0.47485,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Cleavage,Embryo,Whole Organism,All anatomical structures
87,DRR032737,DRX029543,DRS049942,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 95 individuals,Dr 32cell 1,SAMD00028134,,sample name:Dr 32cell 1|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:32cell|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028134,DRX029543,Dr 32cell 1,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028134,,,,3870906500.0,38709065.0,DRR032737,0:100 1:0,A:1030407751;C:909948718;G:905608620;T:1024897443;N:43968,100,0,,,1030407751,909948718,905608620,1024897443,43968,DRX029543,DRS049942,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.93364,,0.02484,,0.77307,,0.47588,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Cleavage,Embryo,Whole Organism,All anatomical structures
88,DRR032736,DRX029542,DRS049941,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 100 individuals,Dr zfs:0000015 2,SAMD00028133,,sample name:Dr zfs:0000015 2|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:zfs:0000015|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028133,DRX029542,Dr zfs:0000015 2,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028133,,,,3129028500.0,31290285.0,DRR032736,0:100 1:0,A:849515903;C:721550282;G:717777586;T:840154982;N:29747,100,0,,,849515903,721550282,717777586,840154982,29747,DRX029542,DRS049941,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92724,,0.07971,,0.74657,,0.47796,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Blastula,Embryo,Whole Organism,All anatomical structures
89,DRR032735,DRX029541,DRS049940,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 100 individuals,Dr zfs:0000015 1,SAMD00028132,,sample name:Dr zfs:0000015 1|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:zfs:0000015|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028132,DRX029541,Dr zfs:0000015 1,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028132,,,,4310219700.0,43102197.0,DRR032735,0:100 1:0,A:1169701983;C:993241399;G:986263558;T:1160969083;N:43677,100,0,,,1169701983,993241399,986263558,1160969083,43677,DRX029541,DRS049940,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92609,,0.07773,,0.74349,,0.47849,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Blastula,Embryo,Whole Organism,All anatomical structures
90,DRR032734,DRX029540,DRS049939,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 107 individuals,Dr 2cell 2,SAMD00028131,,sample name:Dr 2cell 2|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:2cell|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028131,DRX029540,Dr 2cell 2,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028131,,,,3687517000.0,36875170.0,DRR032734,0:100 1:0,A:975272080;C:873518282;G:869743434;T:968941851;N:41353,100,0,,,975272080,873518282,869743434,968941851,41353,DRX029540,DRS049939,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.93204,,0.02088,,0.81639,,0.47553,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Cleavage,Embryo,Whole Organism,All anatomical structures
91,DRR032733,DRX029539,DRS049938,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 108 individuals,Dr 2cell 1,SAMD00028130,,sample name:Dr 2cell 1|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:2cell|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028130,DRX029539,Dr 2cell 1,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028130,,,,4156651100.0,41566511.0,DRR032733,0:100 1:0,A:1099943617;C:985498415;G:978884426;T:1092278665;N:45977,100,0,,,1099943617,985498415,978884426,1092278665,45977,DRX029539,DRS049938,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.93452,,0.02198,,0.81197,,0.47342,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Cleavage,Embryo,Whole Organism,All anatomical structures
92,DRR032732,DRX029538,DRS049937,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 80 individuals,Dr 14somite 3,SAMD00028129,,sample name:Dr 14somite 3|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:14somite|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028129,DRX029538,Dr 14somite 3,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028129,,,,3734610500.0,37346105.0,DRR032732,0:100 1:0,A:1009418541;C:863710067;G:858061383;T:1003378800;N:41709,100,0,,,1009418541,863710067,858061383,1003378800,41709,DRX029538,DRS049937,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92401,,0.08815,,0.70816,,0.46602,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Segmentation,Embryo,Whole Organism,All anatomical structures
93,DRR032731,DRX029537,DRS049936,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 80 individuals,Dr 14somite 2,SAMD00028128,,sample name:Dr 14somite 2|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:14somite|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028128,DRX029537,Dr 14somite 2,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028128,,,,3715174200.0,37151742.0,DRR032731,0:100 1:0,A:1000703508;C:862290629;G:858173468;T:993968396;N:38199,100,0,,,1000703508,862290629,858173468,993968396,38199,DRX029537,DRS049936,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92491,,0.0819,,0.71068,,0.46957,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Segmentation,Embryo,Whole Organism,All anatomical structures
94,DRR032730,DRX029536,DRS049935,DRP003810,PRJDB3785,EXPANDE project,DRP003810,Other,EXPression AloNg Development and Evolution EXPANDE project aims to identify gene expression profiles expanded during embryogenesis and evolution. In brief  taking advantages of Illumina sequencing  RNAseq profiles of early to late embryos of 8 chordate species were identified with biological replicates two or more biological replicates.,,,mRNA extracted from pooled embryos of 80 individuals,Dr 14somite 1,SAMD00028127,,sample name:Dr 14somite 1|strain:Riken WT Wild Type|tissue type:whole embryo|dev stage:14somite|genotype:wild type|phenotype:wild type|sex:male  female  and mixed,,,,,,,,,Illumina HiSeq 2000 sequencing of SAMD00028127,DRX029536,Dr 14somite 1,1,Total RNA QIAGEN RNeasy followed by TruSeq,,RNA-Seq,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>100</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,DRP003810,Illumina HiSeq 2000 sequencing of SAMD00028127,,,,3744386000.0,37443860.0,DRR032730,0:100 1:0,A:1014537326;C:864070910;G:859190201;T:1006549502;N:38061,100,0,,,1014537326,864070910,859190201,1006549502,38061,DRX029536,DRS049935,DRA003460,"UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo","UT-BS|Lab for embryology, Department of Biological Sciences, University of Tokyo",1,0.92378,,0.08957,,0.7068,,0.47493,,100,,B,,usable mapping rate,illumina,hiseq_era,unknown,other,trueseq,bulk,unknown,unknown,,Japan,2017-09-20,Segmentation,Embryo,Whole Organism,All anatomical structures
7946,ERR015563,ERX005934,ERS012707,ERP000263,PRJEB2208,Zebrafish gene three prime end pull down for genome annotation,E-MTAB-308,Transcriptome Analysis,,,,,E MTAB 308:Zebrafish embryo 2 dpf 2,SAMEA898403,Wellcome Sanger Institute,Age:2 days|Alias:E MTAB 308:Zebrafish embryo 2 dpf 2|Broker name:ArrayExpress|Description:Protocols: Zebrafish embyos or tissues were collected from a Tuebingen strain incross and grown at 28 C. Collected samples were snap frozen on dry ice and stored at  70 C Total RNA was extracted using Trizol Reagent Invitrogen following the manufacturer's instructions. Pellets were resuspended RNase free 10 mM Tris pH 7.5 and the RNA was quantified using a NanoDrop ND 1000 Spectrophotometer Axon Instruments.|DevelopmentalStage:embryo|INSDC center alias:SC|INSDC center name:Wellcome Sanger Institute|INSDC first public:2010 08 19T15:57:35Z|INSDC last update:2018 03 08T15:25:04Z|INSDC status:public|InitialTimePoint:fertilization|OrganismPart:whole organism|SRA accession:ERS012707|Sample Name:ERS012707|Sex:unknown sex|StrainOrLine:Tuebingen|Title:Danio rerio,,,,,,,,,Illumina Genome Analyzer II paired end sequencing; Zebrafish gene 3 prime end pull down for genome annotation,E MTAB 308:Illumina Genome Analyzer II sequencing of Zebrafish embryo 2 dpf three prime pull down paired end 250 to 300 bp insert,Zebrafish embro 2 dpf mRNA three prime end,Zebrafish gene three prime end pull down for genome annotation,20 ug of total RNA was fragmented using RNA Fragmentation Reagent Ambion for 5 minutes at 70 C and ethanol precipitated with glycogen and LiCl.   RNA was annealed to the oligo stBPM1polyT22 biotin GGCCAGTCCTGGAGTTTTTTTTTTTTTTTTTTTTTTVN and bound to streptavidin  magnetic beads.   post washing by pull down on a magnet  the bound RNA was reverse transcribed with SuperScript II Invitrogen and a second strand synthesised with DNA polymerase I Promega and RNase H NEB.   post further washing  the double strand cDNA was released from the beads with BpmI NEB.   The cDNA was recovered with the QIAgen PCR Purification Kit and made into a standard Illumina library following the manufacturer's protocol with a fragment size of 250 to 300 bp.,Experimental Factor: AGE:2 d|Experimental Factor: DEVELOPMENTAL STAGE:embryo|Experimental Factor: ORGANISM PART:whole organism|Experimental Factor: SEX:unknown sex,RNA-Seq,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina Genome Analyzer II,,ERP000263,Illumina Genome Analyzer II paired end sequencing; Zebrafish gene three prime end pull down for genome annotation,ENA FIRST PUBLIC:2010 08 19|ENA LAST UPDATE:2018 11 16,3444_2.srf,srf,990308120.0,6515185.0,E MTAB 308:Illumina Genome Analyzer II sequencing of Zebrafish embryo 2 dpf three prime pull down paired end 250 to 300 bp insert,0:76 1:76,A:279665076;C:200433201;G:189692111;T:304366697;N:16151035,76,76,,,279665076,200433201,189692111,304366697,16151035,ERX005934,ERS012707,ERA010603,SC|Wellcome Trust Sanger Institute,SC|Wellcome Trust Sanger Institute,2,0.93964,0.94008,0.40095,0.39969,0.74424,0.74915,0.49535,0.49761,76,76,B,B,biological fallback assumption,illumina,early_illumina,3prime,other,unknown,bulk,unknown,unknown,,United Kingdom,2010-08-19,Hatching,Embryo,Whole Organism,All anatomical structures
7947,ERR015564,ERX005933,ERS012706,ERP000263,PRJEB2208,Zebrafish gene three prime end pull down for genome annotation,E-MTAB-308,Transcriptome Analysis,,,,,E MTAB 308:Zebrafish embryo 3 dpf 2,SAMEA898404,Wellcome Sanger Institute,Age:3 days|Alias:E MTAB 308:Zebrafish embryo 3 dpf 2|Broker name:ArrayExpress|Description:Protocols: Zebrafish embyos or tissues were collected from a Tuebingen strain incross and grown at 28 C. Collected samples were snap frozen on dry ice and stored at  70 C Total RNA was extracted using Trizol Reagent Invitrogen following the manufacturer's instructions. Pellets were resuspended RNase free 10 mM Tris pH 7.5 and the RNA was quantified using a NanoDrop ND 1000 Spectrophotometer Axon Instruments.|DevelopmentalStage:embryo|INSDC center alias:SC|INSDC center name:Wellcome Sanger Institute|INSDC first public:2010 08 19T15:57:35Z|INSDC last update:2018 03 08T15:25:04Z|INSDC status:public|InitialTimePoint:fertilization|OrganismPart:whole organism|SRA accession:ERS012706|Sample Name:ERS012706|Sex:unknown sex|StrainOrLine:Tuebingen|Title:Danio rerio,,,,,,,,,Illumina Genome Analyzer II paired end sequencing; Zebrafish gene 3 prime end pull down for genome annotation,E MTAB 308:Illumina Genome Analyzer II sequencing of Zebrafish embryo 3 dpf three prime pull down paired end 250 to 300 bp insert,Zebrafish embro 3 dpf mRNA three prime end,Zebrafish gene three prime end pull down for genome annotation,20 ug of total RNA was fragmented using RNA Fragmentation Reagent Ambion for 5 minutes at 70 C and ethanol precipitated with glycogen and LiCl.   RNA was annealed to the oligo stBPM1polyT22 biotin GGCCAGTCCTGGAGTTTTTTTTTTTTTTTTTTTTTTVN and bound to streptavidin  magnetic beads.   post washing by pull down on a magnet  the bound RNA was reverse transcribed with SuperScript II Invitrogen and a second strand synthesised with DNA polymerase I Promega and RNase H NEB.   post further washing  the double strand cDNA was released from the beads with BpmI NEB.   The cDNA was recovered with the QIAgen PCR Purification Kit and made into a standard Illumina library following the manufacturer's protocol with a fragment size of 250 to 300 bp.,Experimental Factor: AGE:3 d|Experimental Factor: DEVELOPMENTAL STAGE:embryo|Experimental Factor: ORGANISM PART:whole organism|Experimental Factor: SEX:unknown sex,RNA-Seq,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina Genome Analyzer II,,ERP000263,Illumina Genome Analyzer II paired end sequencing; Zebrafish gene three prime end pull down for genome annotation,ENA FIRST PUBLIC:2010 08 19|ENA LAST UPDATE:2018 11 16,3444_3.srf,srf,1572215648.0,10343524.0,E MTAB 308:Illumina Genome Analyzer II sequencing of Zebrafish embryo 3 dpf three prime pull down paired end 250 to 300 bp insert,0:76 1:76,A:395189954;C:374281517;G:356666372;T:420372888;N:25704917,76,76,,,395189954,374281517,356666372,420372888,25704917,ERX005933,ERS012706,ERA010603,SC|Wellcome Trust Sanger Institute,SC|Wellcome Trust Sanger Institute,2,0.96337,0.96626,0.12558,0.12974,0.7824,0.79086,0.40731,0.41802,76,76,B,B,biological fallback assumption,illumina,early_illumina,3prime,other,unknown,bulk,unknown,unknown,,United Kingdom,2010-08-19,Larval,Larval,Whole Organism,All anatomical structures
7948,ERR015562,ERX005932,ERS012705,ERP000263,PRJEB2208,Zebrafish gene three prime end pull down for genome annotation,E-MTAB-308,Transcriptome Analysis,,,,,E MTAB 308:Zebrafish embryo 1 dpf 2,SAMEA898401,Wellcome Sanger Institute,Age:1 days|Alias:E MTAB 308:Zebrafish embryo 1 dpf 2|Broker name:ArrayExpress|Description:Protocols: Zebrafish embyos or tissues were collected from a Tuebingen strain incross and grown at 28 C. Collected samples were snap frozen on dry ice and stored at  70 C Total RNA was extracted using Trizol Reagent Invitrogen following the manufacturer's instructions. Pellets were resuspended RNase free 10 mM Tris pH 7.5 and the RNA was quantified using a NanoDrop ND 1000 Spectrophotometer Axon Instruments.|DevelopmentalStage:embryo|INSDC center alias:SC|INSDC center name:Wellcome Sanger Institute|INSDC first public:2010 08 19T15:57:35Z|INSDC last update:2018 03 08T15:25:04Z|INSDC status:public|InitialTimePoint:fertilization|OrganismPart:whole organism|SRA accession:ERS012705|Sample Name:ERS012705|Sex:unknown sex|StrainOrLine:Tuebingen|Title:Danio rerio,,,,,,,,,Illumina Genome Analyzer II paired end sequencing; Zebrafish gene 3 prime end pull down for genome annotation,E MTAB 308:Illumina Genome Analyzer II sequencing of Zebrafish embryo 1 dpf three prime pull down paired end 250 to 300 bp insert,Zebrafish embro 1 dpf mRNA three prime end,Zebrafish gene three prime end pull down for genome annotation,20 ug of total RNA was fragmented using RNA Fragmentation Reagent Ambion for 5 minutes at 70 C and ethanol precipitated with glycogen and LiCl.   RNA was annealed to the oligo stBPM1polyT22 biotin GGCCAGTCCTGGAGTTTTTTTTTTTTTTTTTTTTTTVN and bound to streptavidin  magnetic beads.   post washing by pull down on a magnet  the bound RNA was reverse transcribed with SuperScript II Invitrogen and a second strand synthesised with DNA polymerase I Promega and RNase H NEB.   post further washing  the double strand cDNA was released from the beads with BpmI NEB.   The cDNA was recovered with the QIAgen PCR Purification Kit and made into a standard Illumina library following the manufacturer's protocol with a fragment size of 250 to 300 bp.,Experimental Factor: AGE:1 d|Experimental Factor: DEVELOPMENTAL STAGE:embryo|Experimental Factor: ORGANISM PART:whole organism|Experimental Factor: SEX:unknown sex,RNA-Seq,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina Genome Analyzer II,,ERP000263,Illumina Genome Analyzer II paired end sequencing; Zebrafish gene three prime end pull down for genome annotation,ENA FIRST PUBLIC:2010 08 19|ENA LAST UPDATE:2018 11 16,3444_1.srf,srf,1358041568.0,8934484.0,E MTAB 308:Illumina Genome Analyzer II sequencing of Zebrafish embryo 1 dpf three prime pull down paired end 250 to 300 bp insert,0:76 1:76,A:376045982;C:281568469;G:271365867;T:407279179;N:21782071,76,76,,,376045982,281568469,271365867,407279179,21782071,ERX005932,ERS012705,ERA010603,SC|Wellcome Trust Sanger Institute,SC|Wellcome Trust Sanger Institute,2,0.94607,0.94542,0.3002,0.30139,0.73584,0.74038,0.51058,0.51233,76,76,B,B,biological fallback assumption,illumina,early_illumina,3prime,other,unknown,bulk,unknown,unknown,,United Kingdom,2010-08-19,Pharyngula,Embryo,Whole Organism,All anatomical structures
7951,ERR015565,ERX005929,ERS012704,ERP000263,PRJEB2208,Zebrafish gene three prime end pull down for genome annotation,E-MTAB-308,Transcriptome Analysis,,,,,E MTAB 308:Zebrafish embryo 5 dpf 2,SAMEA980815,SC,Age:5 days|Alias:E MTAB 308:Zebrafish embryo 5 dpf 2|Broker name:ArrayExpress|Description:Protocols: Zebrafish embyos or tissues were collected from a Tuebingen strain incross and grown at 28 C. Collected samples were snap frozen on dry ice and stored at  70 C Total RNA was extracted using Trizol Reagent Invitrogen following the manufacturer's instructions. Pellets were resuspended RNase free 10 mM Tris pH 7.5 and the RNA was quantified using a NanoDrop ND 1000 Spectrophotometer Axon Instruments.|DevelopmentalStage:embryo|INSDC center name:SC|INSDC first public:2010 08 19T15:57:35Z|INSDC last update:2018 03 08T15:25:04Z|INSDC status:public|InitialTimePoint:fertilization|OrganismPart:whole organism|SRA accession:ERS012704|Sample Name:ERS012704|Sex:unknown sex|StrainOrLine:Tuebingen|Title:Danio rerio,,,,,,,,,Illumina Genome Analyzer II paired end sequencing; Zebrafish gene 3 prime end pull down for genome annotation,E MTAB 308:Illumina Genome Analyzer II sequencing of Zebrafish embryo 5 dpf three prime pull down paired end 250 to 300 bp insert,Zebrafish embro 5 dpf mRNA three prime end,Zebrafish gene three prime end pull down for genome annotation,20 ug of total RNA was fragmented using RNA Fragmentation Reagent Ambion for 5 minutes at 70 C and ethanol precipitated with glycogen and LiCl.   RNA was annealed to the oligo stBPM1polyT22 biotin GGCCAGTCCTGGAGTTTTTTTTTTTTTTTTTTTTTTVN and bound to streptavidin  magnetic beads.   post washing by pull down on a magnet  the bound RNA was reverse transcribed with SuperScript II Invitrogen and a second strand synthesised with DNA polymerase I Promega and RNase H NEB.   post further washing  the double strand cDNA was released from the beads with BpmI NEB.   The cDNA was recovered with the QIAgen PCR Purification Kit and made into a standard Illumina library following the manufacturer's protocol with a fragment size of 250 to 300 bp.,Experimental Factor: AGE:5 d|Experimental Factor: DEVELOPMENTAL STAGE:embryo|Experimental Factor: ORGANISM PART:whole organism|Experimental Factor: SEX:unknown sex,RNA-Seq,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina Genome Analyzer II,,ERP000263,Illumina Genome Analyzer II paired end sequencing; Zebrafish gene three prime end pull down for genome annotation,ENA FIRST PUBLIC:2010 08 19|ENA LAST UPDATE:2018 11 16,3444_5.srf,srf,1399629832.0,9208091.0,E MTAB 308:Illumina Genome Analyzer II sequencing of Zebrafish embryo 5 dpf three prime pull down paired end 250 to 300 bp insert,0:76 1:76,A:389123080;C:293319124;G:279736391;T:414876632;N:22574605,76,76,,,389123080,293319124,279736391,414876632,22574605,ERX005929,ERS012704,ERA010603,SC|Wellcome Trust Sanger Institute,SC|Wellcome Trust Sanger Institute,2,0.92999,0.9332,0.45309,0.46162,0.72419,0.73919,0.49892,0.50512,76,76,B,B,biological fallback assumption,illumina,early_illumina,3prime,other,unknown,bulk,unknown,unknown,,United Kingdom,2010-08-19,Larval,Larval,Whole Organism,All anatomical structures
7952,ERR015568,ERX005928,ERS000087,ERP000263,PRJEB2208,Zebrafish gene three prime end pull down for genome annotation,E-MTAB-308,Transcriptome Analysis,,,,,ZF male sample1,SAMEA708829,Wellcome Sanger Institute,Alias:ZF male sample1|Description:RNA extracted from whole male adult zebrafish without xxx|INSDC center alias:SC|INSDC center name:Wellcome Sanger Institute|INSDC first public:2010 02 26T10:44:13Z|INSDC last update:2018 03 08T15:24:37Z|INSDC status:public|SRA accession:ERS000087|Sample Name:ERS000087|Sex:male|Strain:Tuebingen|Title:Danio rerio,,,,,,,,,Illumina Genome Analyzer II paired end sequencing; Zebrafish gene 3 prime end pull down for genome annotation,E MTAB 308:Illumina Genome Analyzer II sequencing of adult Zebrafish male body dpf three prime pull down paired end 250 to 300 bp insert,Zebrafish adult male body mRNA three prime end,Zebrafish gene three prime end pull down for genome annotation,20 ug of total RNA was fragmented using RNA Fragmentation Reagent Ambion for 5 minutes at 70 C and ethanol precipitated with glycogen and LiCl.   RNA was annealed to the oligo stBPM1polyT22 biotin GGCCAGTCCTGGAGTTTTTTTTTTTTTTTTTTTTTTVN and bound to streptavidin  magnetic beads.   post washing by pull down on a magnet  the bound RNA was reverse transcribed with SuperScript II Invitrogen and a second strand synthesised with DNA polymerase I Promega and RNase H NEB.   post further washing  the double strand cDNA was released from the beads with BpmI NEB.   The cDNA was recovered with the QIAgen PCR Purification Kit and made into a standard Illumina library following the manufacturer's protocol with a fragment size of 250 to 300 bp.,Experimental Factor: DEVELOPMENTAL STAGE:adult|Experimental Factor: ORGANISM PART:whole fish without xxx|Experimental Factor: SEX:male,RNA-Seq,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina Genome Analyzer II,,ERP000263,Illumina Genome Analyzer II paired end sequencing; Zebrafish gene three prime end pull down for genome annotation,ENA FIRST PUBLIC:2010 08 19|ENA LAST UPDATE:2018 11 16,3444_8.srf,srf,1012667320.0,6662285.0,E MTAB 308:Illumina Genome Analyzer II sequencing of adult Zebrafish male body dpf three prime pull down paired end 250 to 300 bp insert,0:76 1:76,A:262408681;C:234183204;G:228254982;T:271015871;N:16804582,76,76,,,262408681,234183204,228254982,271015871,16804582,ERX005928,ERS000087,ERA010603,SC|Wellcome Trust Sanger Institute,SC|Wellcome Trust Sanger Institute,2,0.95123,0.96116,0.23163,0.22276,0.77847,0.78648,0.42301,0.43528,76,76,B,B,biological fallback assumption,illumina,early_illumina,3prime,other,unknown,bulk,unknown,unknown,,United Kingdom,2010-02-26,Adult,Adult,Whole Organism,All anatomical structures
25295,SRR25764045,SRX21486723,SRS18719024,SRP457105,PRJNA1009809,Dynamics of the zebrafish tRNAome during the maternal to zygotic transition [Ribo Seq],GSE241753,Other,Time course analysis of tRNA abundance during zebrafish early embryonic development. Overall design: Wild type TLAB strain zebrafish embryos were grown in standard housing conditions.Unfertilized eggs 0 hpf were collected or embryos were staged and collected at consecutive developmental time points  namelyat the 256 cell 2.5 hpf  1000 cell 3 hpf  sphere 4 hpf  shield 6 hpf and bud 10 hpf stages. Eggs and embryos were either flash frozen in liquid nitrogen or immediately processed. Samples were used for western blotting analysis  polysome profiling  ribosome profiling or mRNA and tRNA sequencing for investigating the regulation of tRNA gene expression and translational status during early zebrafish embryogenesis.,parent bioproject:PRJNA1009800,pubmed:39402326,,WT bud 10 hpf Ribo seq rep1,GSM7734770,,source name:Gastrula|strain:TLAB strain|tissue:Gastrula|developmental stage:Bud 10 hpf|treatment:100 µg/ml CHX  100 µg/ml TIG|geo loc name:missing|collection date:missing,WT bud 10 hpf Ribo seq rep1,We identified the A site location in each mapped read with Scikit ribo [Fang et al. 2018]  which uses a random forest with recursive feature selection and a generalized linear model for accurate A site prediction based on matched ribosome profiling and RNA Seq datasets. Kallisto 0.44.0 with parameters  b 100   single  l 180  s 20  t 40 was used to quantify transcript abundances in Transcripts Per Million TPM from RNA Seq data based on the reference set of MANE annotated transcripts see Codon usage analysis for description of this annotation. To avoid memory errors due to the large size of the human genome and the presence of multiple transcript isoforms  all RNAfold dependencies in Scikit ribo were omitted  and the index was built separately for each chromosome. To make the hg38 GTF compatible with Scikit ribo  transcript/UTR annotations were removed. For each transcript  the start codon in the first exon and the stop codon in the last exon were adjusted to represent transcript start and end coordinates  taking into account the gene strand. To estimate codon dwell times  short 20 23 nt and long 28 33 nt ribosome footprints were analyzed separately. rRNA filtered reads were aligned to GRCz11.108 using STAR v2.6.1c [Dobin et al.  2013] the following options:   outFilterMultimapNmax 1   seedSearchStartLmax 15   outSAMtype BAM SortedByCoordinate   outFilterMismatchNmax 2   alignEndsType EndToEnd   quantMode TranscriptomeSAM   outSAMattributes NH HI AS nM NM MD We identified the A site location in each mapped read with Scikit ribo [Fang et al. 2018]  which uses a random forest with recursive feature selection and a generalized linear model for accurate A site prediction based on matched ribosome profiling and RNA Seq datasets. Kallisto 0.44.0 with parameters  b 100   single  l 180  s 20  t 40 was used to quantify transcript abundances in Transcripts Per Million TPM from RNA Seq data based on the reference set of MANE annotated transcripts see Codon usage analysis for description of this annotation. To avoid memory errors due to the large size of the human genome and the presence of multiple transcript isoforms  all RNAfold dependencies in Scikit ribo were omitted  and the index was built separately for each chromosome. To make the GRCz11 GTF compatible with Scikit ribo  transcript/UTR annotations were removed. For each transcript  the start codon in the first exon and the stop codon in the last exon were adjusted to represent transcript start and end coordinates  taking into account the gene strand. To estimate codon dwell times  we utilised ribosome footprints with length 30 34 nt. Assembly: GRCz11 Supplementary files format and content: csv; codon dwell times from scikit ribo for all samples Supplementary files format and content: csv; transcript per million TPM counts per transcript in MANE annotation Library strategy: Ribo Seq,Gastrula,unperturbed growth conditions in E3 medium for zebrafish embryos.,200 whole embryos were flash frozen in liquid nitrogen and subsequently lysed in footprint lysis buffer containing 100 µg/ml CHX and 100 µg/ml TIG  0.1% NP 40  10 µg/ml aprotinin  20 µM leupeptin  2.5 µM pepstatin A  0.5 mM AEBSF and 1x Phosphatase Inhibitor Cocktail. Samples were vortexed vigorously  triturated through a 26G gauge needle  and spun down for 7 minutes at 16 000xg/ 4°C. Supernatant was transferred to a new tube. 20 µg RNA in 200 µl polysome lysis buffer were digested with 50 U RNase I for 45 minutes at 2 000 rpm/22°C. post incubation on ice for 5 minutes  extracts were pre cleared by centrifugation for 5 minutes at 3 000 g/ 4°C. Ribosomes were pelleted through 3 ml of a sucrose cushion 1 M sucrose  20 mM Tris pH=8.0  140 mM KCl  5 mM MgCl2  1 mM DTT by spinning the layered solutions in the Type 70 Ti rotor for 120 minutes at 50 000 rpm/ 4°C. Ribosome pellets were rinsed once  dissolved in 200 µl drug free polysome lysis buffer  and incubated with 200 U hiPSC or 300 U NPC RNase I for 45 minutes at 2 000 rpm/22°C. Ribosome footprint libraries were prepared essentially as described McGlincy and Ingolia  2017; Wu 2019 with minor modifications. RNase I digestion was stopped by addition of 100 U Superase In and extracts were loaded on a sucrose cushion. The pellet was dissolved in 400 µl LiDS/LET lysis buffer and RNA was extracted with the acid phenol protocol. Fragments in the range of 19 to 32 nucleotides were isolated by gel size selection  with T4 PNK and ligated to pre adenylated adapters containing 5 random nucleotides at their 5’ ends McGlinzy 2017 with T4 RNA Ligase 2  truncated KQ. Adapter ligated RNA was subjected to rRNA depletion using the Ribo Seq riboPOOL h/m/r depletion kit siTOOLs for CHX only samples  and legacy RiboZero Gold kit Illumina for CHX+TIG samples The rRNA depleted footprints were reverse transcribed with Protoscript II and cDNA was circularized with recombinant TS2126 RNA ligase 1 commercially available as CircLigase. Libraries were constructed from circularized cDNA with KAPA HiFi DNA Polymerase and single end sequencing was performed on a NextSeq 500 platform Illumina.,Embryos were grown in standard housing conditions namely28°C at a 14/10 hour light/dark cycle.,strain:TLAB strain|tissue:Gastrula|developmental stage:Bud 10 hpf|treatment:100 µg/ml CHX  100 µg/ml TIG,GSM7734770,GSM7734770: WT bud 10 hpf Ribo seq rep1; Danio rerio; OTHER,GSM7734770 r1,GSM7734770,1,200 whole embryos were flash frozen in liquid nitrogen and subsequently lysed in footprint lysis buffer containing 100 µg/ml CHX and 100 µg/ml TIG  0.1% NP 40  10 µg/ml aprotinin  20 µM leupeptin  2.5 µM pepstatin A  0.5 mM AEBSF and 1x Phosphatase Inhibitor Cocktail. Samples were vortexed vigorously  triturated through a 26G gauge needle  and spun down for 7 minutes at 16 000xg/ 4°C. Supernatant was transferred to a new tube. 20 µg RNA in 200 µl polysome lysis buffer were digested with 50 U RNase I for 45 minutes at 2 000 rpm/22°C. post incubation on ice for 5 minutes  extracts were pre cleared by centrifugation for 5 minutes at 3 000 g/ 4°C. Ribosomes were pelleted through 3 ml of a sucrose cushion 1 M sucrose  20 mM Tris pH=8.0  140 mM KCl  5 mM MgCl2  1 mM DTT by spinning the layered solutions in the Type 70 Ti rotor for 120 minutes at 50 000 rpm/ 4°C. Ribosome pellets were rinsed once  dissolved in 200 µl drug free polysome lysis buffer  and incubated with 200 U hiPSC or 300 U NPC RNase I for 45 minutes at 2 000 rpm/22°C. Ribosome footprint libraries were prepared essentially as described McGlincy and Ingolia  2017; Wu 2019 with minor modifications. RNase I digestion was stopped by addition of 100 U Superase In and extracts were loaded on a sucrose cushion. The pellet was dissolved in 400 µl LiDS/LET lysis buffer and RNA was extracted with the acid phenol protocol. Fragments in the range of 19 to 32 nucleotides were isolated by gel size selection  with T4 PNK and ligated to pre adenylated adapters containing 5 random nucleotides at their five prime ends McGlinzy 2017 with T4 RNA Ligase 2  truncated KQ. Adapter ligated RNA was subjected to rRNA depletion using the Ribo Seq riboPOOL h/m/r depletion kit siTOOLs for CHX only samples  and legacy RiboZero Gold kit Illumina for CHX+TIG samples The rRNA depleted footprints were reverse transcribed with Protoscript II and cDNA was circularized with recombinant TS2126 RNA ligase 1 commercially available as CircLigase. Libraries were constructed from circularized cDNA with KAPA HiFi DNA Polymerase and single end sequencing was performed on a NextSeq 500 platform Illumina.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,NextSeq 500,,SRP457105,,,WT_ribo_bud_1.fastq.gz,fastq,1476985703.0,55745002.0,GSM7734770 r1,0:26.50,A:266369385;C:466461341;G:473469631;T:270671671;N:13675,26,,,,266369385,466461341,473469631,270671671,13675,SRX21486723,SRS18719024,SRA1700409,"Mechanisms of Protein Biogenesis, Max Planck Institute for Biochemistry",Max Planck Institute of Biochemistry,1,0.773,,0.14121,,0.82242,,0.78464,,30,,B,,usable mapping rate,illumina,nextseq,5prime,rrna_depletion,ribozero,bulk,unknown,unknown,,Germany,2023-08-28,Gastrula,Embryo,Whole Organism,All anatomical structures
25296,SRR25764046,SRX21486722,SRS18719023,SRP457105,PRJNA1009809,Dynamics of the zebrafish tRNAome during the maternal to zygotic transition [Ribo Seq],GSE241753,Other,Time course analysis of tRNA abundance during zebrafish early embryonic development. Overall design: Wild type TLAB strain zebrafish embryos were grown in standard housing conditions.Unfertilized eggs 0 hpf were collected or embryos were staged and collected at consecutive developmental time points  namelyat the 256 cell 2.5 hpf  1000 cell 3 hpf  sphere 4 hpf  shield 6 hpf and bud 10 hpf stages. Eggs and embryos were either flash frozen in liquid nitrogen or immediately processed. Samples were used for western blotting analysis  polysome profiling  ribosome profiling or mRNA and tRNA sequencing for investigating the regulation of tRNA gene expression and translational status during early zebrafish embryogenesis.,parent bioproject:PRJNA1009800,pubmed:39402326,,WT sphere 4 hpf Ribo seq rep1,GSM7734769,,source name:Blastula|strain:TLAB strain|tissue:Blastula|developmental stage:Sphere 4 hpf|treatment:100 µg/ml CHX  100 µg/ml TIG|geo loc name:missing|collection date:missing,WT sphere 4 hpf Ribo seq rep1,We identified the A site location in each mapped read with Scikit ribo [Fang et al. 2018]  which uses a random forest with recursive feature selection and a generalized linear model for accurate A site prediction based on matched ribosome profiling and RNA Seq datasets. Kallisto 0.44.0 with parameters  b 100   single  l 180  s 20  t 40 was used to quantify transcript abundances in Transcripts Per Million TPM from RNA Seq data based on the reference set of MANE annotated transcripts see Codon usage analysis for description of this annotation. To avoid memory errors due to the large size of the human genome and the presence of multiple transcript isoforms  all RNAfold dependencies in Scikit ribo were omitted  and the index was built separately for each chromosome. To make the hg38 GTF compatible with Scikit ribo  transcript/UTR annotations were removed. For each transcript  the start codon in the first exon and the stop codon in the last exon were adjusted to represent transcript start and end coordinates  taking into account the gene strand. To estimate codon dwell times  short 20 23 nt and long 28 33 nt ribosome footprints were analyzed separately. rRNA filtered reads were aligned to GRCz11.108 using STAR v2.6.1c [Dobin et al.  2013] the following options:   outFilterMultimapNmax 1   seedSearchStartLmax 15   outSAMtype BAM SortedByCoordinate   outFilterMismatchNmax 2   alignEndsType EndToEnd   quantMode TranscriptomeSAM   outSAMattributes NH HI AS nM NM MD We identified the A site location in each mapped read with Scikit ribo [Fang et al. 2018]  which uses a random forest with recursive feature selection and a generalized linear model for accurate A site prediction based on matched ribosome profiling and RNA Seq datasets. Kallisto 0.44.0 with parameters  b 100   single  l 180  s 20  t 40 was used to quantify transcript abundances in Transcripts Per Million TPM from RNA Seq data based on the reference set of MANE annotated transcripts see Codon usage analysis for description of this annotation. To avoid memory errors due to the large size of the human genome and the presence of multiple transcript isoforms  all RNAfold dependencies in Scikit ribo were omitted  and the index was built separately for each chromosome. To make the GRCz11 GTF compatible with Scikit ribo  transcript/UTR annotations were removed. For each transcript  the start codon in the first exon and the stop codon in the last exon were adjusted to represent transcript start and end coordinates  taking into account the gene strand. To estimate codon dwell times  we utilised ribosome footprints with length 30 34 nt. Assembly: GRCz11 Supplementary files format and content: csv; codon dwell times from scikit ribo for all samples Supplementary files format and content: csv; transcript per million TPM counts per transcript in MANE annotation Library strategy: Ribo Seq,Blastula,unperturbed growth conditions in E3 medium for zebrafish embryos.,200 whole embryos were flash frozen in liquid nitrogen and subsequently lysed in footprint lysis buffer containing 100 µg/ml CHX and 100 µg/ml TIG  0.1% NP 40  10 µg/ml aprotinin  20 µM leupeptin  2.5 µM pepstatin A  0.5 mM AEBSF and 1x Phosphatase Inhibitor Cocktail. Samples were vortexed vigorously  triturated through a 26G gauge needle  and spun down for 7 minutes at 16 000xg/ 4°C. Supernatant was transferred to a new tube. 20 µg RNA in 200 µl polysome lysis buffer were digested with 50 U RNase I for 45 minutes at 2 000 rpm/22°C. post incubation on ice for 5 minutes  extracts were pre cleared by centrifugation for 5 minutes at 3 000 g/ 4°C. Ribosomes were pelleted through 3 ml of a sucrose cushion 1 M sucrose  20 mM Tris pH=8.0  140 mM KCl  5 mM MgCl2  1 mM DTT by spinning the layered solutions in the Type 70 Ti rotor for 120 minutes at 50 000 rpm/ 4°C. Ribosome pellets were rinsed once  dissolved in 200 µl drug free polysome lysis buffer  and incubated with 200 U hiPSC or 300 U NPC RNase I for 45 minutes at 2 000 rpm/22°C. Ribosome footprint libraries were prepared essentially as described McGlincy and Ingolia  2017; Wu 2019 with minor modifications. RNase I digestion was stopped by addition of 100 U Superase In and extracts were loaded on a sucrose cushion. The pellet was dissolved in 400 µl LiDS/LET lysis buffer and RNA was extracted with the acid phenol protocol. Fragments in the range of 19 to 32 nucleotides were isolated by gel size selection  with T4 PNK and ligated to pre adenylated adapters containing 5 random nucleotides at their 5’ ends McGlinzy 2017 with T4 RNA Ligase 2  truncated KQ. Adapter ligated RNA was subjected to rRNA depletion using the Ribo Seq riboPOOL h/m/r depletion kit siTOOLs for CHX only samples  and legacy RiboZero Gold kit Illumina for CHX+TIG samples The rRNA depleted footprints were reverse transcribed with Protoscript II and cDNA was circularized with recombinant TS2126 RNA ligase 1 commercially available as CircLigase. Libraries were constructed from circularized cDNA with KAPA HiFi DNA Polymerase and single end sequencing was performed on a NextSeq 500 platform Illumina.,Embryos were grown in standard housing conditions namely28°C at a 14/10 hour light/dark cycle.,strain:TLAB strain|tissue:Blastula|developmental stage:Sphere 4 hpf|treatment:100 µg/ml CHX  100 µg/ml TIG,GSM7734769,GSM7734769: WT sphere 4 hpf Ribo seq rep1; Danio rerio; OTHER,GSM7734769 r1,GSM7734769,1,200 whole embryos were flash frozen in liquid nitrogen and subsequently lysed in footprint lysis buffer containing 100 µg/ml CHX and 100 µg/ml TIG  0.1% NP 40  10 µg/ml aprotinin  20 µM leupeptin  2.5 µM pepstatin A  0.5 mM AEBSF and 1x Phosphatase Inhibitor Cocktail. Samples were vortexed vigorously  triturated through a 26G gauge needle  and spun down for 7 minutes at 16 000xg/ 4°C. Supernatant was transferred to a new tube. 20 µg RNA in 200 µl polysome lysis buffer were digested with 50 U RNase I for 45 minutes at 2 000 rpm/22°C. post incubation on ice for 5 minutes  extracts were pre cleared by centrifugation for 5 minutes at 3 000 g/ 4°C. Ribosomes were pelleted through 3 ml of a sucrose cushion 1 M sucrose  20 mM Tris pH=8.0  140 mM KCl  5 mM MgCl2  1 mM DTT by spinning the layered solutions in the Type 70 Ti rotor for 120 minutes at 50 000 rpm/ 4°C. Ribosome pellets were rinsed once  dissolved in 200 µl drug free polysome lysis buffer  and incubated with 200 U hiPSC or 300 U NPC RNase I for 45 minutes at 2 000 rpm/22°C. Ribosome footprint libraries were prepared essentially as described McGlincy and Ingolia  2017; Wu 2019 with minor modifications. RNase I digestion was stopped by addition of 100 U Superase In and extracts were loaded on a sucrose cushion. The pellet was dissolved in 400 µl LiDS/LET lysis buffer and RNA was extracted with the acid phenol protocol. Fragments in the range of 19 to 32 nucleotides were isolated by gel size selection  with T4 PNK and ligated to pre adenylated adapters containing 5 random nucleotides at their five prime ends McGlinzy 2017 with T4 RNA Ligase 2  truncated KQ. Adapter ligated RNA was subjected to rRNA depletion using the Ribo Seq riboPOOL h/m/r depletion kit siTOOLs for CHX only samples  and legacy RiboZero Gold kit Illumina for CHX+TIG samples The rRNA depleted footprints were reverse transcribed with Protoscript II and cDNA was circularized with recombinant TS2126 RNA ligase 1 commercially available as CircLigase. Libraries were constructed from circularized cDNA with KAPA HiFi DNA Polymerase and single end sequencing was performed on a NextSeq 500 platform Illumina.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,NextSeq 500,,SRP457105,,,WT_ribo_sphere_1.fastq.gz,fastq,1336027898.0,47907512.0,GSM7734769 r1,0:27.89,A:229038284;C:439455963;G:429428292;T:238088512;N:16847,27,,,,229038284,439455963,429428292,238088512,16847,SRX21486722,SRS18719023,SRA1700409,"Mechanisms of Protein Biogenesis, Max Planck Institute for Biochemistry",Max Planck Institute of Biochemistry,1,0.85867,,0.20418,,0.8776,,0.79481,,24,,B,,usable mapping rate,illumina,nextseq,5prime,rrna_depletion,ribozero,bulk,unknown,unknown,,Germany,2023-08-28,Blastula,Embryo,Whole Organism,All anatomical structures
31943,SRR28790250,SRX24354554,SRS21112322,SRP503769,PRJNA1104180,Double stranded RNA triggers a distinct integrated stress response in the early embryo [RNA seq  Ribo seq],GSE265771,Other,Double stranded RNA dsRNA is associated with virus infections and is present as by products during the transcription of synthetic mRNA  which has been widely used in gene gain of function studies and serves as a core component in emerging mRNA based therapies1 4. The presence of dsRNA in host cells induces an integrated stress response that functions to prevent virus replication and infection5 6. Unlike differentiated cells  undifferentiated cells adopt a distinct defense strategy against RNA virus infection7  but the mechanism is unclear. We show a previously unidentified response triggered by dsRNA in the early embryo. Although dsRNA causes a global protein translation inhibition in a PKR eIF2a independent manner and leads to developmental delay and cell necrosis  it also strongly induces p53 activation  which then upregulates Interferon Stimulated Genes independently of interferon ligands. Importantly  we demonstrate that the burst of p53 signaling dose not result in cell death but functions as a protective mechanism against deleterious translation blockage by slowing down global protein degradation via ISGylation. Our work has identified a distinct dsRNA induced stress response in the embryo  reflecting an ancient innate immune memory before the establishment of the IFN system. It also raises the provocative question as to the original protective role of p53 during evolution. Overall design: To characterize the distribution of ribosomes on mRNA  we performed Ribosome profiling Ribo seq analysis with RNA seq on dsRNA injected embryos.,,,,Zebrafish  Riboseq  high dose  dsRNA2,GSM8228804,,source name:whole embryo|tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:dsRNA|geo loc name:missing|collection date:missing,Zebrafish  Riboseq  high dose  dsRNA2,The library underwent quality control assessment  and was subjected to Illumina Novaseq 6000 sequencing. Bio informatics analysis of RIBOseq profilling was performed to analyze the library data. Assembly: GRCz10 Supplementary files format and content: Excel file includes raw counts for each Sample Library strategy: Ribo seq,whole embryo,,Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components  including undigested material such as cell nuclei.Then  RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA  while the RNA fragments protected by ribosomes were preserved. Subsequently  an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation  and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx  and the corresponding bands were collected. The recovered bands were subjected to ligation reaction  where a preadenylylated and 3’ blocked linker 5’ rApp CTGTAGGCACCATCAAT NH2 3’ was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently  the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre  CL4111K  resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA  a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR  and barcode sequences were incorporated.,,tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:dsRNA,GSM8228804,GSM8228804: Zebrafish  Riboseq  high dose  dsRNA2; Danio rerio; OTHER,GSM8228804 r1,GSM8228804,1,Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components  including undigested material such as cell nuclei.Then  RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA  while the RNA fragments protected by ribosomes were preserved. Subsequently  an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation  and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx  and the corresponding bands were collected. The recovered bands were subjected to ligation reaction  where a preadenylylated and three prime blocked linker five prime rApp CTGTAGGCACCATCAAT NH2 three prime was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently  the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre  CL4111K  resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA  a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR  and barcode sequences were incorporated.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP503769,,,dsR_2.R1.raw.fastq.gz dsR_2.R2.raw.fastq.gz,fastq fastq,15877367026.0,52574063.0,GSM8228804 r1,0:151 1:151,A:3017944891;C:2902425937;G:7224440180;T:2731881397;N:674621,151,151,,,3017944891,2902425937,7224440180,2731881397,674621,SRX24354554,SRS21112322,SRA1852128,ShanDong University,ShanDong University,,,,,,,,,,,,T,T,mates < 9% mapping rate,illumina,novaseq_era,3prime,random_priming,unknown,bulk,unknown,unknown,,China,2024-04-24,Gastrula,Embryo,Whole Organism,All anatomical structures
31944,SRR28790251,SRX24354553,SRS21112321,SRP503769,PRJNA1104180,Double stranded RNA triggers a distinct integrated stress response in the early embryo [RNA seq  Ribo seq],GSE265771,Other,Double stranded RNA dsRNA is associated with virus infections and is present as by products during the transcription of synthetic mRNA  which has been widely used in gene gain of function studies and serves as a core component in emerging mRNA based therapies1 4. The presence of dsRNA in host cells induces an integrated stress response that functions to prevent virus replication and infection5 6. Unlike differentiated cells  undifferentiated cells adopt a distinct defense strategy against RNA virus infection7  but the mechanism is unclear. We show a previously unidentified response triggered by dsRNA in the early embryo. Although dsRNA causes a global protein translation inhibition in a PKR eIF2a independent manner and leads to developmental delay and cell necrosis  it also strongly induces p53 activation  which then upregulates Interferon Stimulated Genes independently of interferon ligands. Importantly  we demonstrate that the burst of p53 signaling dose not result in cell death but functions as a protective mechanism against deleterious translation blockage by slowing down global protein degradation via ISGylation. Our work has identified a distinct dsRNA induced stress response in the embryo  reflecting an ancient innate immune memory before the establishment of the IFN system. It also raises the provocative question as to the original protective role of p53 during evolution. Overall design: To characterize the distribution of ribosomes on mRNA  we performed Ribosome profiling Ribo seq analysis with RNA seq on dsRNA injected embryos.,,,,Zebrafish  Riboseq  high dose  dsRNA1,GSM8228803,,source name:whole embryo|tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:dsRNA|geo loc name:missing|collection date:missing,Zebrafish  Riboseq  high dose  dsRNA1,The library underwent quality control assessment  and was subjected to Illumina Novaseq 6000 sequencing. Bio informatics analysis of RIBOseq profilling was performed to analyze the library data. Assembly: GRCz10 Supplementary files format and content: Excel file includes raw counts for each Sample Library strategy: Ribo seq,whole embryo,,Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components  including undigested material such as cell nuclei.Then  RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA  while the RNA fragments protected by ribosomes were preserved. Subsequently  an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation  and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx  and the corresponding bands were collected. The recovered bands were subjected to ligation reaction  where a preadenylylated and 3’ blocked linker 5’ rApp CTGTAGGCACCATCAAT NH2 3’ was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently  the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre  CL4111K  resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA  a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR  and barcode sequences were incorporated.,,tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:dsRNA,GSM8228803,GSM8228803: Zebrafish  Riboseq  high dose  dsRNA1; Danio rerio; OTHER,GSM8228803 r1,GSM8228803,1,Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components  including undigested material such as cell nuclei.Then  RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA  while the RNA fragments protected by ribosomes were preserved. Subsequently  an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation  and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx  and the corresponding bands were collected. The recovered bands were subjected to ligation reaction  where a preadenylylated and three prime blocked linker five prime rApp CTGTAGGCACCATCAAT NH2 three prime was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently  the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre  CL4111K  resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA  a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR  and barcode sequences were incorporated.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP503769,,,dsR_1.R1.raw.fastq.gz dsR_1.R2.raw.fastq.gz,fastq fastq,13027928774.0,43138837.0,GSM8228803 r1,0:151 1:151,A:2538099201;C:2400262316;G:5852047273;T:2236961902;N:558082,151,151,,,2538099201,2400262316,5852047273,2236961902,558082,SRX24354553,SRS21112321,SRA1852128,ShanDong University,ShanDong University,,,,,,,,,,,,T,T,mates < 9% mapping rate,illumina,novaseq_era,3prime,random_priming,unknown,bulk,unknown,unknown,,China,2024-04-24,Gastrula,Embryo,Whole Organism,All anatomical structures
31945,SRR28790252,SRX24354552,SRS21112320,SRP503769,PRJNA1104180,Double stranded RNA triggers a distinct integrated stress response in the early embryo [RNA seq  Ribo seq],GSE265771,Other,Double stranded RNA dsRNA is associated with virus infections and is present as by products during the transcription of synthetic mRNA  which has been widely used in gene gain of function studies and serves as a core component in emerging mRNA based therapies1 4. The presence of dsRNA in host cells induces an integrated stress response that functions to prevent virus replication and infection5 6. Unlike differentiated cells  undifferentiated cells adopt a distinct defense strategy against RNA virus infection7  but the mechanism is unclear. We show a previously unidentified response triggered by dsRNA in the early embryo. Although dsRNA causes a global protein translation inhibition in a PKR eIF2a independent manner and leads to developmental delay and cell necrosis  it also strongly induces p53 activation  which then upregulates Interferon Stimulated Genes independently of interferon ligands. Importantly  we demonstrate that the burst of p53 signaling dose not result in cell death but functions as a protective mechanism against deleterious translation blockage by slowing down global protein degradation via ISGylation. Our work has identified a distinct dsRNA induced stress response in the embryo  reflecting an ancient innate immune memory before the establishment of the IFN system. It also raises the provocative question as to the original protective role of p53 during evolution. Overall design: To characterize the distribution of ribosomes on mRNA  we performed Ribosome profiling Ribo seq analysis with RNA seq on dsRNA injected embryos.,,,,Zebrafish  Riboseq  high dose  uninj2,GSM8228802,,source name:whole embryo|tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:uninjected|geo loc name:missing|collection date:missing,Zebrafish  Riboseq  high dose  uninj2,The library underwent quality control assessment  and was subjected to Illumina Novaseq 6000 sequencing. Bio informatics analysis of RIBOseq profilling was performed to analyze the library data. Assembly: GRCz10 Supplementary files format and content: Excel file includes raw counts for each Sample Library strategy: Ribo seq,whole embryo,,Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components  including undigested material such as cell nuclei.Then  RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA  while the RNA fragments protected by ribosomes were preserved. Subsequently  an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation  and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx  and the corresponding bands were collected. The recovered bands were subjected to ligation reaction  where a preadenylylated and 3’ blocked linker 5’ rApp CTGTAGGCACCATCAAT NH2 3’ was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently  the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre  CL4111K  resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA  a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR  and barcode sequences were incorporated.,,tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:uninjected,GSM8228802,GSM8228802: Zebrafish  Riboseq  high dose  uninj2; Danio rerio; OTHER,GSM8228802 r1,GSM8228802,1,Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components  including undigested material such as cell nuclei.Then  RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA  while the RNA fragments protected by ribosomes were preserved. Subsequently  an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation  and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx  and the corresponding bands were collected. The recovered bands were subjected to ligation reaction  where a preadenylylated and three prime blocked linker five prime rApp CTGTAGGCACCATCAAT NH2 three prime was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently  the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre  CL4111K  resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA  a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR  and barcode sequences were incorporated.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP503769,,,un_2.R1.raw.fastq.gz un_2.R2.raw.fastq.gz,fastq fastq,13637060056.0,45155828.0,GSM8228802 r1,0:151 1:151,A:2596015831;C:2388366938;G:6201010197;T:2451085273;N:581817,151,151,,,2596015831,2388366938,6201010197,2451085273,581817,SRX24354552,SRS21112320,SRA1852128,ShanDong University,ShanDong University,,,,,,,,,,,,T,T,mates < 9% mapping rate,illumina,novaseq_era,3prime,random_priming,unknown,bulk,unknown,unknown,,China,2024-04-24,Gastrula,Embryo,Whole Organism,All anatomical structures
31946,SRR28790253,SRX24354551,SRS21112319,SRP503769,PRJNA1104180,Double stranded RNA triggers a distinct integrated stress response in the early embryo [RNA seq  Ribo seq],GSE265771,Other,Double stranded RNA dsRNA is associated with virus infections and is present as by products during the transcription of synthetic mRNA  which has been widely used in gene gain of function studies and serves as a core component in emerging mRNA based therapies1 4. The presence of dsRNA in host cells induces an integrated stress response that functions to prevent virus replication and infection5 6. Unlike differentiated cells  undifferentiated cells adopt a distinct defense strategy against RNA virus infection7  but the mechanism is unclear. We show a previously unidentified response triggered by dsRNA in the early embryo. Although dsRNA causes a global protein translation inhibition in a PKR eIF2a independent manner and leads to developmental delay and cell necrosis  it also strongly induces p53 activation  which then upregulates Interferon Stimulated Genes independently of interferon ligands. Importantly  we demonstrate that the burst of p53 signaling dose not result in cell death but functions as a protective mechanism against deleterious translation blockage by slowing down global protein degradation via ISGylation. Our work has identified a distinct dsRNA induced stress response in the embryo  reflecting an ancient innate immune memory before the establishment of the IFN system. It also raises the provocative question as to the original protective role of p53 during evolution. Overall design: To characterize the distribution of ribosomes on mRNA  we performed Ribosome profiling Ribo seq analysis with RNA seq on dsRNA injected embryos.,,,,Zebrafish  Riboseq  high dose  uninj1,GSM8228801,,source name:whole embryo|tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:uninjected|geo loc name:missing|collection date:missing,Zebrafish  Riboseq  high dose  uninj1,The library underwent quality control assessment  and was subjected to Illumina Novaseq 6000 sequencing. Bio informatics analysis of RIBOseq profilling was performed to analyze the library data. Assembly: GRCz10 Supplementary files format and content: Excel file includes raw counts for each Sample Library strategy: Ribo seq,whole embryo,,Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components  including undigested material such as cell nuclei.Then  RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA  while the RNA fragments protected by ribosomes were preserved. Subsequently  an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation  and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx  and the corresponding bands were collected. The recovered bands were subjected to ligation reaction  where a preadenylylated and 3’ blocked linker 5’ rApp CTGTAGGCACCATCAAT NH2 3’ was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently  the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre  CL4111K  resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA  a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR  and barcode sequences were incorporated.,,tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:uninjected,GSM8228801,GSM8228801: Zebrafish  Riboseq  high dose  uninj1; Danio rerio; OTHER,GSM8228801 r1,GSM8228801,1,Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components  including undigested material such as cell nuclei.Then  RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA  while the RNA fragments protected by ribosomes were preserved. Subsequently  an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation  and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx  and the corresponding bands were collected. The recovered bands were subjected to ligation reaction  where a preadenylylated and three prime blocked linker five prime rApp CTGTAGGCACCATCAAT NH2 three prime was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently  the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre  CL4111K  resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA  a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR  and barcode sequences were incorporated.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP503769,,,un_1.R1.raw.fastq.gz un_1.R2.raw.fastq.gz,fastq fastq,13499333560.0,44699780.0,GSM8228801 r1,0:151 1:151,A:2587725963;C:2439789499;G:6080777505;T:2390460459;N:580134,151,151,,,2587725963,2439789499,6080777505,2390460459,580134,SRX24354551,SRS21112319,SRA1852128,ShanDong University,ShanDong University,,,,,,,,,,,,T,T,mates < 9% mapping rate,illumina,novaseq_era,3prime,random_priming,unknown,bulk,unknown,unknown,,China,2024-04-24,Gastrula,Embryo,Whole Organism,All anatomical structures
31947,SRR28790254,SRX24354550,SRS21112318,SRP503769,PRJNA1104180,Double stranded RNA triggers a distinct integrated stress response in the early embryo [RNA seq  Ribo seq],GSE265771,Other,Double stranded RNA dsRNA is associated with virus infections and is present as by products during the transcription of synthetic mRNA  which has been widely used in gene gain of function studies and serves as a core component in emerging mRNA based therapies1 4. The presence of dsRNA in host cells induces an integrated stress response that functions to prevent virus replication and infection5 6. Unlike differentiated cells  undifferentiated cells adopt a distinct defense strategy against RNA virus infection7  but the mechanism is unclear. We show a previously unidentified response triggered by dsRNA in the early embryo. Although dsRNA causes a global protein translation inhibition in a PKR eIF2a independent manner and leads to developmental delay and cell necrosis  it also strongly induces p53 activation  which then upregulates Interferon Stimulated Genes independently of interferon ligands. Importantly  we demonstrate that the burst of p53 signaling dose not result in cell death but functions as a protective mechanism against deleterious translation blockage by slowing down global protein degradation via ISGylation. Our work has identified a distinct dsRNA induced stress response in the embryo  reflecting an ancient innate immune memory before the establishment of the IFN system. It also raises the provocative question as to the original protective role of p53 during evolution. Overall design: To characterize the distribution of ribosomes on mRNA  we performed Ribosome profiling Ribo seq analysis with RNA seq on dsRNA injected embryos.,,,,Zebrafish  Riboseq  low dose  dsRNA2,GSM8228800,,source name:whole embryo|tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:dsRNA|geo loc name:missing|collection date:missing,Zebrafish  Riboseq  low dose  dsRNA2,The library underwent quality control assessment  and was subjected to Illumina Novaseq 6000 sequencing. Bio informatics analysis of RIBOseq profilling was performed to analyze the library data. Assembly: GRCz10 Supplementary files format and content: Excel file includes raw counts for each Sample Library strategy: Ribo seq,whole embryo,,Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components  including undigested material such as cell nuclei.Then  RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA  while the RNA fragments protected by ribosomes were preserved. Subsequently  an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation  and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx  and the corresponding bands were collected. The recovered bands were subjected to ligation reaction  where a preadenylylated and 3’ blocked linker 5’ rApp CTGTAGGCACCATCAAT NH2 3’ was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently  the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre  CL4111K  resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA  a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR  and barcode sequences were incorporated.,,tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:dsRNA,GSM8228800,GSM8228800: Zebrafish  Riboseq  low dose  dsRNA2; Danio rerio; OTHER,GSM8228800 r1,GSM8228800,1,Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components  including undigested material such as cell nuclei.Then  RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA  while the RNA fragments protected by ribosomes were preserved. Subsequently  an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation  and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx  and the corresponding bands were collected. The recovered bands were subjected to ligation reaction  where a preadenylylated and three prime blocked linker five prime rApp CTGTAGGCACCATCAAT NH2 three prime was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently  the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre  CL4111K  resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA  a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR  and barcode sequences were incorporated.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP503769,,,dsR_2.raw.1.fastq.gz dsR_2.raw.2.fastq.gz,fastq fastq,13335544766.0,44157433.0,GSM8228800 r1,0:151 1:151,A:2383528724;C:2262795235;G:6593680813;T:2094730217;N:809777,151,151,,,2383528724,2262795235,6593680813,2094730217,809777,SRX24354550,SRS21112318,SRA1852128,ShanDong University,ShanDong University,,,,,,,,,,,,T,T,mates < 9% mapping rate,illumina,novaseq_era,3prime,random_priming,unknown,bulk,unknown,unknown,,China,2024-04-24,Gastrula,Embryo,Whole Organism,All anatomical structures
31948,SRR28790255,SRX24354549,SRS21112317,SRP503769,PRJNA1104180,Double stranded RNA triggers a distinct integrated stress response in the early embryo [RNA seq  Ribo seq],GSE265771,Other,Double stranded RNA dsRNA is associated with virus infections and is present as by products during the transcription of synthetic mRNA  which has been widely used in gene gain of function studies and serves as a core component in emerging mRNA based therapies1 4. The presence of dsRNA in host cells induces an integrated stress response that functions to prevent virus replication and infection5 6. Unlike differentiated cells  undifferentiated cells adopt a distinct defense strategy against RNA virus infection7  but the mechanism is unclear. We show a previously unidentified response triggered by dsRNA in the early embryo. Although dsRNA causes a global protein translation inhibition in a PKR eIF2a independent manner and leads to developmental delay and cell necrosis  it also strongly induces p53 activation  which then upregulates Interferon Stimulated Genes independently of interferon ligands. Importantly  we demonstrate that the burst of p53 signaling dose not result in cell death but functions as a protective mechanism against deleterious translation blockage by slowing down global protein degradation via ISGylation. Our work has identified a distinct dsRNA induced stress response in the embryo  reflecting an ancient innate immune memory before the establishment of the IFN system. It also raises the provocative question as to the original protective role of p53 during evolution. Overall design: To characterize the distribution of ribosomes on mRNA  we performed Ribosome profiling Ribo seq analysis with RNA seq on dsRNA injected embryos.,,,,Zebrafish  Riboseq  low dose  dsRNA1,GSM8228799,,source name:whole embryo|tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:dsRNA|geo loc name:missing|collection date:missing,Zebrafish  Riboseq  low dose  dsRNA1,The library underwent quality control assessment  and was subjected to Illumina Novaseq 6000 sequencing. Bio informatics analysis of RIBOseq profilling was performed to analyze the library data. Assembly: GRCz10 Supplementary files format and content: Excel file includes raw counts for each Sample Library strategy: Ribo seq,whole embryo,,Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components  including undigested material such as cell nuclei.Then  RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA  while the RNA fragments protected by ribosomes were preserved. Subsequently  an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation  and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx  and the corresponding bands were collected. The recovered bands were subjected to ligation reaction  where a preadenylylated and 3’ blocked linker 5’ rApp CTGTAGGCACCATCAAT NH2 3’ was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently  the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre  CL4111K  resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA  a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR  and barcode sequences were incorporated.,,tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:dsRNA,GSM8228799,GSM8228799: Zebrafish  Riboseq  low dose  dsRNA1; Danio rerio; OTHER,GSM8228799 r1,GSM8228799,1,Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components  including undigested material such as cell nuclei.Then  RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA  while the RNA fragments protected by ribosomes were preserved. Subsequently  an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation  and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx  and the corresponding bands were collected. The recovered bands were subjected to ligation reaction  where a preadenylylated and three prime blocked linker five prime rApp CTGTAGGCACCATCAAT NH2 three prime was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently  the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre  CL4111K  resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA  a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR  and barcode sequences were incorporated.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP503769,,,dsR_1.raw.1.fastq.gz dsR_1.raw.2.fastq.gz,fastq fastq,12530433302.0,41491501.0,GSM8228799 r1,0:151 1:151,A:2300207705;C:2164355626;G:6050556625;T:2014558360;N:754986,151,151,,,2300207705,2164355626,6050556625,2014558360,754986,SRX24354549,SRS21112317,SRA1852128,ShanDong University,ShanDong University,,,,,,,,,,,,T,T,mates < 9% mapping rate,illumina,novaseq_era,3prime,random_priming,unknown,bulk,unknown,unknown,,China,2024-04-24,Gastrula,Embryo,Whole Organism,All anatomical structures
31949,SRR28790256,SRX24354548,SRS21112316,SRP503769,PRJNA1104180,Double stranded RNA triggers a distinct integrated stress response in the early embryo [RNA seq  Ribo seq],GSE265771,Other,Double stranded RNA dsRNA is associated with virus infections and is present as by products during the transcription of synthetic mRNA  which has been widely used in gene gain of function studies and serves as a core component in emerging mRNA based therapies1 4. The presence of dsRNA in host cells induces an integrated stress response that functions to prevent virus replication and infection5 6. Unlike differentiated cells  undifferentiated cells adopt a distinct defense strategy against RNA virus infection7  but the mechanism is unclear. We show a previously unidentified response triggered by dsRNA in the early embryo. Although dsRNA causes a global protein translation inhibition in a PKR eIF2a independent manner and leads to developmental delay and cell necrosis  it also strongly induces p53 activation  which then upregulates Interferon Stimulated Genes independently of interferon ligands. Importantly  we demonstrate that the burst of p53 signaling dose not result in cell death but functions as a protective mechanism against deleterious translation blockage by slowing down global protein degradation via ISGylation. Our work has identified a distinct dsRNA induced stress response in the embryo  reflecting an ancient innate immune memory before the establishment of the IFN system. It also raises the provocative question as to the original protective role of p53 during evolution. Overall design: To characterize the distribution of ribosomes on mRNA  we performed Ribosome profiling Ribo seq analysis with RNA seq on dsRNA injected embryos.,,,,Zebrafish  Riboseq  low dose  uninj2,GSM8228798,,source name:whole embryo|tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:uninjected|geo loc name:missing|collection date:missing,Zebrafish  Riboseq  low dose  uninj2,The library underwent quality control assessment  and was subjected to Illumina Novaseq 6000 sequencing. Bio informatics analysis of RIBOseq profilling was performed to analyze the library data. Assembly: GRCz10 Supplementary files format and content: Excel file includes raw counts for each Sample Library strategy: Ribo seq,whole embryo,,Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components  including undigested material such as cell nuclei.Then  RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA  while the RNA fragments protected by ribosomes were preserved. Subsequently  an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation  and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx  and the corresponding bands were collected. The recovered bands were subjected to ligation reaction  where a preadenylylated and 3’ blocked linker 5’ rApp CTGTAGGCACCATCAAT NH2 3’ was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently  the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre  CL4111K  resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA  a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR  and barcode sequences were incorporated.,,tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:uninjected,GSM8228798,GSM8228798: Zebrafish  Riboseq  low dose  uninj2; Danio rerio; OTHER,GSM8228798 r1,GSM8228798,1,Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components  including undigested material such as cell nuclei.Then  RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA  while the RNA fragments protected by ribosomes were preserved. Subsequently  an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation  and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx  and the corresponding bands were collected. The recovered bands were subjected to ligation reaction  where a preadenylylated and three prime blocked linker five prime rApp CTGTAGGCACCATCAAT NH2 three prime was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently  the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre  CL4111K  resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA  a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR  and barcode sequences were incorporated.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP503769,,,un_2.raw.1.fastq.gz un_2.raw.2.fastq.gz,fastq fastq,12132018896.0,40172248.0,GSM8228798 r1,0:151 1:151,A:2225485331;C:2027579165;G:5953072417;T:1925141784;N:740199,151,151,,,2225485331,2027579165,5953072417,1925141784,740199,SRX24354548,SRS21112316,SRA1852128,ShanDong University,ShanDong University,,,,,,,,,,,,T,T,mates < 9% mapping rate,illumina,novaseq_era,3prime,random_priming,unknown,bulk,unknown,unknown,,China,2024-04-24,Gastrula,Embryo,Whole Organism,All anatomical structures
31950,SRR28790257,SRX24354547,SRS21112315,SRP503769,PRJNA1104180,Double stranded RNA triggers a distinct integrated stress response in the early embryo [RNA seq  Ribo seq],GSE265771,Other,Double stranded RNA dsRNA is associated with virus infections and is present as by products during the transcription of synthetic mRNA  which has been widely used in gene gain of function studies and serves as a core component in emerging mRNA based therapies1 4. The presence of dsRNA in host cells induces an integrated stress response that functions to prevent virus replication and infection5 6. Unlike differentiated cells  undifferentiated cells adopt a distinct defense strategy against RNA virus infection7  but the mechanism is unclear. We show a previously unidentified response triggered by dsRNA in the early embryo. Although dsRNA causes a global protein translation inhibition in a PKR eIF2a independent manner and leads to developmental delay and cell necrosis  it also strongly induces p53 activation  which then upregulates Interferon Stimulated Genes independently of interferon ligands. Importantly  we demonstrate that the burst of p53 signaling dose not result in cell death but functions as a protective mechanism against deleterious translation blockage by slowing down global protein degradation via ISGylation. Our work has identified a distinct dsRNA induced stress response in the embryo  reflecting an ancient innate immune memory before the establishment of the IFN system. It also raises the provocative question as to the original protective role of p53 during evolution. Overall design: To characterize the distribution of ribosomes on mRNA  we performed Ribosome profiling Ribo seq analysis with RNA seq on dsRNA injected embryos.,,,,Zebrafish  Riboseq  low dose  uninj1,GSM8228797,,source name:whole embryo|tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:uninjected|geo loc name:missing|collection date:missing,Zebrafish  Riboseq  low dose  uninj1,The library underwent quality control assessment  and was subjected to Illumina Novaseq 6000 sequencing. Bio informatics analysis of RIBOseq profilling was performed to analyze the library data. Assembly: GRCz10 Supplementary files format and content: Excel file includes raw counts for each Sample Library strategy: Ribo seq,whole embryo,,Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components  including undigested material such as cell nuclei.Then  RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA  while the RNA fragments protected by ribosomes were preserved. Subsequently  an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation  and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx  and the corresponding bands were collected. The recovered bands were subjected to ligation reaction  where a preadenylylated and 3’ blocked linker 5’ rApp CTGTAGGCACCATCAAT NH2 3’ was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently  the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre  CL4111K  resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA  a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR  and barcode sequences were incorporated.,,tissue:whole embryo|developmental stage:50% epiboly stage|cell type:embryonic cell|genotype:wild type|treatment:uninjected,GSM8228797,GSM8228797: Zebrafish  Riboseq  low dose  uninj1; Danio rerio; OTHER,GSM8228797 r1,GSM8228797,1,Both control group and dsRNA injected embryos at xxx hpf were thoroughly lysed on ice using lysis buffer.The lysate was then centrifuged at 12000g at 4°C for 10 minutes to remove cellular components  including undigested material such as cell nuclei.Then  RNaseI was added to the supernatant and incubated at 25°C for 30 minutes to digest RNA  while the RNA fragments protected by ribosomes were preserved. Subsequently  an RNA inhibitor was added to terminate the digestion reaction. The lysate was subjected to sucrose density gradient centrifugation  and fractions containing single peaks at 260nm wavelength around the 80S monosome region were collected. The collected mixture was then thoroughly lysed using Trizol for RNA extraction. RNA fragments within the size range of 26 34nt was separated using polyacrylamide gel electrophoresis without xxx  and the corresponding bands were collected. The recovered bands were subjected to ligation reaction  where a preadenylylated and three prime blocked linker five prime rApp CTGTAGGCACCATCAAT NH2 three prime was attached to the three prime end of the RNA. The ligated products were then recovered by performing polyacrylamide gel electrophoresis. Subsequently  the recovered RNA fragments were reverse transcribed using a reverse primer to obtain extended cDNA molecules. The purified cDNA was subjected to circularization by using CircLigase Epicentre  CL4111K  resulting in the formation of circular cDNA molecules. To remove residual rRNA components in the cDNA  a method involving hybridization with complementary primers specific to rRNA and subsequent heat denaturation was employed. The remaining circularized cDNA was then amplified through PCR  and barcode sequences were incorporated.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP503769,,,un_1.raw.1.fastq.gz un_1.raw.2.fastq.gz,fastq fastq,10423454802.0,34514751.0,GSM8228797 r1,0:151 1:151,A:1931094519;C:1788894999;G:5028533789;T:1674294221;N:637274,151,151,,,1931094519,1788894999,5028533789,1674294221,637274,SRX24354547,SRS21112315,SRA1852128,ShanDong University,ShanDong University,,,,,,,,,,,,T,T,mates < 9% mapping rate,illumina,novaseq_era,3prime,random_priming,unknown,bulk,unknown,unknown,,China,2024-04-24,Gastrula,Embryo,Whole Organism,All anatomical structures
34614,SRR32137213,SRX27483377,SRS23906281,SRP560121,PRJNA1216205,rbm24a is an organizer component of germ plasm to determine germ cell fate 202409 validation,GSE288109,Other,The  formation of germ cells is a critical issue for the continuation of species. A large group of animals follow a preformation strategy to generate their primordial germ cells PGCs. They produce a set of localized maternal mRNAs and proteins to form phase separated germ plasm that functions as the PGC determinants  but the mechanisms underlying the assembly of germ plasm is poorly understood. This study identifies Rbm24a as an enssential localized germ plasm protein component that controls the formation of large and functional germ plasm granules. Rbm24a is complexed with Buc and interacts with germ plasm mRNAs  which determines the specific grasp of germ plasm mRNAs into the phase separated aggregates. Rbm24a absent granules fail to undergo kinesin dependent transport towards the cleavage furrows  where small particles fuse into large ones. The loss of maternal rbm24a causes the complete degradation of germ plasm components and the disappearance of PGCs  resulting in totally sterile animals. Our work establishes that Rbm24 is a critical nucleating organizer component of germ plasm  highlighting an emerging common mechanism to read and recruit RNA component into phase separated condensates. Overall design: To comprehensively characterize the RNA binding landscape of Rbm24a in zebrafish  we performed RIP seq analysis using rbm24a GFP knock in samples  with IgG serving as the control.,,,,Zebrafish  RIP seq  Rbm24a Knock in2 IP2,GSM8759069,,source name:whole embryo|tissue:whole embryo|cell line:4 cell stage|cell type:embryonic cell|genotype:knock in|geo loc name:missing|collection date:missing,Zebrafish  RIP seq  Rbm24a Knock in2 IP2,Each library was amplified by a 15 cycle PCR  and 150 bp paired end sequencing was subjected to Illumina Nova seq 6000 to obtain the raw data. Assembly: GRCz11 Supplementary files format and content: tab delimited text file includes raw counts for each Sample Supplementary files format and content: Bigwig file for each Sample,whole embryo,,Total RNAs were extracted from Mrbm24a embryos at 4 cell sphere 24 hpf stage  using TRIzol reagent Invitrogen  and were precipitated by cold isopropanol 50%  v/v in the presence of carrier glycogen 20 µg/each sample. 1 μg of immunoprecipitated RNA was utilized for library construction. Ribosomal RNA depletion was performed using RNase H based probes followed by magnetic bead purification. RNA fragmentation was achieved through magnesium catalyzed hydrolysis under elevated temperature 94°C for 6 min. First strand cDNA synthesis was initiated with random hexamer primers and reverse transcriptase  followed by second strand synthesis using DNA polymerase I with dUTP incorporation for strand marking. Double stranded cDNA fragments underwent end repair and three prime adenylation using a single tube enzymatic reaction system. Illumina compatible adaptors with unique dual indexes were ligated to both ends of the DNA fragments. Size selection of adaptor ligated products 180 250 bp was performed using double sided SPRI bead purification. PCR amplification was conducted with P5/P7 primers for 12 cycles  followed by AMPure XP bead purification to remove primer dimers. Library quality was verified by Agilent 2100 Bioanalyzer DNA High Sensitivity chip analysis. Indexed libraries were pooled in equimolar ratios and subjected to paired end sequencing 2×150 bp on an Illumina NovaSeq 6000 platform following the manufacturer's standardized workflow.,Zebrafish embryos were maintained in 1/3x Ringer's at 28.5C,tissue:whole embryo|cell line:4 cell stage|cell type:embryonic cell|genotype:knock in,GSM8759069,GSM8759069: Zebrafish  RIP seq  Rbm24a Knock in2 IP2; Danio rerio; RIP Seq,GSM8759069 r1,GSM8759069,1,Total RNAs were extracted from Mrbm24a embryos at 4 cell sphere 24 hpf stage  using TRIzol reagent Invitrogen  and were precipitated by cold isopropanol 50%  v/v in the presence of carrier glycogen 20 µg/each sample. 1 μg of immunoprecipitated RNA was utilized for library construction. Ribosomal RNA depletion was performed using RNase H based probes followed by magnetic bead purification. RNA fragmentation was achieved through magnesium catalyzed hydrolysis under elevated temperature 94°C for 6 min. First strand cDNA synthesis was initiated with random hexamer primers and reverse transcriptase  followed by second strand synthesis using DNA polymerase I with dUTP incorporation for strand marking. Double stranded cDNA fragments underwent end repair and three prime adenylation using a single tube enzymatic reaction system. Illumina compatible adaptors with unique dual indexes were ligated to both ends of the DNA fragments. Size selection of adaptor ligated products 180 250 bp was performed using double sided SPRI bead purification. PCR amplification was conducted with P5/P7 primers for 12 cycles  followed by AMPure XP bead purification to remove primer dimers. Library quality was verified by Agilent 2100 Bioanalyzer DNA High Sensitivity chip analysis. Indexed libraries were pooled in equimolar ratios and subjected to paired end sequencing 2×150 bp on an Illumina NovaSeq 6000 platform following the manufacturer's standardized workflow.,,RIP-Seq,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP560121,,,RIP_IP2.R1.fq.gz RIP_IP2.R2.fq.gz,fastq fastq,18215080774.0,60314837.0,GSM8759069 r1,0:151 1:151,A:3824315591;C:5035112770;G:5644328190;T:3707941000;N:3383223,151,151,,,3824315591,5035112770,5644328190,3707941000,3383223,SRX27483377,SRS23906281,SRA2061743,"Ang Li, College of Life Sciences, shandong university","Ang Li, College of Life Sciences, shandong university",,,,,,,,,,,,B,B,mate2-mate1 similar by mapping diff,illumina,novaseq_era,unknown,random_priming,unknown,bulk,unknown,unknown,,China,2025-01-27,Multi-stage,Embryo,Whole Organism,All anatomical structures
34615,SRR32137214,SRX27483376,SRS23906280,SRP560121,PRJNA1216205,rbm24a is an organizer component of germ plasm to determine germ cell fate 202409 validation,GSE288109,Other,The  formation of germ cells is a critical issue for the continuation of species. A large group of animals follow a preformation strategy to generate their primordial germ cells PGCs. They produce a set of localized maternal mRNAs and proteins to form phase separated germ plasm that functions as the PGC determinants  but the mechanisms underlying the assembly of germ plasm is poorly understood. This study identifies Rbm24a as an enssential localized germ plasm protein component that controls the formation of large and functional germ plasm granules. Rbm24a is complexed with Buc and interacts with germ plasm mRNAs  which determines the specific grasp of germ plasm mRNAs into the phase separated aggregates. Rbm24a absent granules fail to undergo kinesin dependent transport towards the cleavage furrows  where small particles fuse into large ones. The loss of maternal rbm24a causes the complete degradation of germ plasm components and the disappearance of PGCs  resulting in totally sterile animals. Our work establishes that Rbm24 is a critical nucleating organizer component of germ plasm  highlighting an emerging common mechanism to read and recruit RNA component into phase separated condensates. Overall design: To comprehensively characterize the RNA binding landscape of Rbm24a in zebrafish  we performed RIP seq analysis using rbm24a GFP knock in samples  with IgG serving as the control.,,,,Zebrafish  RIP seq  Rbm24a Knock in1 IP1,GSM8759068,,source name:whole embryo|tissue:whole embryo|cell line:4 cell stage|cell type:embryonic cell|genotype:knock in|geo loc name:missing|collection date:missing,Zebrafish  RIP seq  Rbm24a Knock in1 IP1,Each library was amplified by a 15 cycle PCR  and 150 bp paired end sequencing was subjected to Illumina Nova seq 6000 to obtain the raw data. Assembly: GRCz11 Supplementary files format and content: tab delimited text file includes raw counts for each Sample Supplementary files format and content: Bigwig file for each Sample,whole embryo,,Total RNAs were extracted from Mrbm24a embryos at 4 cell sphere 24 hpf stage  using TRIzol reagent Invitrogen  and were precipitated by cold isopropanol 50%  v/v in the presence of carrier glycogen 20 µg/each sample. 1 μg of immunoprecipitated RNA was utilized for library construction. Ribosomal RNA depletion was performed using RNase H based probes followed by magnetic bead purification. RNA fragmentation was achieved through magnesium catalyzed hydrolysis under elevated temperature 94°C for 6 min. First strand cDNA synthesis was initiated with random hexamer primers and reverse transcriptase  followed by second strand synthesis using DNA polymerase I with dUTP incorporation for strand marking. Double stranded cDNA fragments underwent end repair and three prime adenylation using a single tube enzymatic reaction system. Illumina compatible adaptors with unique dual indexes were ligated to both ends of the DNA fragments. Size selection of adaptor ligated products 180 250 bp was performed using double sided SPRI bead purification. PCR amplification was conducted with P5/P7 primers for 12 cycles  followed by AMPure XP bead purification to remove primer dimers. Library quality was verified by Agilent 2100 Bioanalyzer DNA High Sensitivity chip analysis. Indexed libraries were pooled in equimolar ratios and subjected to paired end sequencing 2×150 bp on an Illumina NovaSeq 6000 platform following the manufacturer's standardized workflow.,Zebrafish embryos were maintained in 1/3x Ringer's at 28.5C,tissue:whole embryo|cell line:4 cell stage|cell type:embryonic cell|genotype:knock in,GSM8759068,GSM8759068: Zebrafish  RIP seq  Rbm24a Knock in1 IP1; Danio rerio; RIP Seq,GSM8759068 r1,GSM8759068,1,Total RNAs were extracted from Mrbm24a embryos at 4 cell sphere 24 hpf stage  using TRIzol reagent Invitrogen  and were precipitated by cold isopropanol 50%  v/v in the presence of carrier glycogen 20 µg/each sample. 1 μg of immunoprecipitated RNA was utilized for library construction. Ribosomal RNA depletion was performed using RNase H based probes followed by magnetic bead purification. RNA fragmentation was achieved through magnesium catalyzed hydrolysis under elevated temperature 94°C for 6 min. First strand cDNA synthesis was initiated with random hexamer primers and reverse transcriptase  followed by second strand synthesis using DNA polymerase I with dUTP incorporation for strand marking. Double stranded cDNA fragments underwent end repair and three prime adenylation using a single tube enzymatic reaction system. Illumina compatible adaptors with unique dual indexes were ligated to both ends of the DNA fragments. Size selection of adaptor ligated products 180 250 bp was performed using double sided SPRI bead purification. PCR amplification was conducted with P5/P7 primers for 12 cycles  followed by AMPure XP bead purification to remove primer dimers. Library quality was verified by Agilent 2100 Bioanalyzer DNA High Sensitivity chip analysis. Indexed libraries were pooled in equimolar ratios and subjected to paired end sequencing 2×150 bp on an Illumina NovaSeq 6000 platform following the manufacturer's standardized workflow.,,RIP-Seq,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP560121,,,RIP_IP1.R1.fq.gz RIP_IP1.R2.fq.gz,fastq fastq,22190472874.0,73478387.0,GSM8759068 r1,0:151 1:151,A:4585155428;C:6233832602;G:6882198251;T:4484977909;N:4308684,151,151,,,4585155428,6233832602,6882198251,4484977909,4308684,SRX27483376,SRS23906280,SRA2061743,"Ang Li, College of Life Sciences, shandong university","Ang Li, College of Life Sciences, shandong university",,,,,,,,,,,,B,B,mate2-mate1 similar by mapping diff,illumina,novaseq_era,unknown,random_priming,unknown,bulk,unknown,unknown,,China,2025-01-27,Multi-stage,Embryo,Whole Organism,All anatomical structures
34616,SRR32137215,SRX27483375,SRS23906279,SRP560121,PRJNA1216205,rbm24a is an organizer component of germ plasm to determine germ cell fate 202409 validation,GSE288109,Other,The  formation of germ cells is a critical issue for the continuation of species. A large group of animals follow a preformation strategy to generate their primordial germ cells PGCs. They produce a set of localized maternal mRNAs and proteins to form phase separated germ plasm that functions as the PGC determinants  but the mechanisms underlying the assembly of germ plasm is poorly understood. This study identifies Rbm24a as an enssential localized germ plasm protein component that controls the formation of large and functional germ plasm granules. Rbm24a is complexed with Buc and interacts with germ plasm mRNAs  which determines the specific grasp of germ plasm mRNAs into the phase separated aggregates. Rbm24a absent granules fail to undergo kinesin dependent transport towards the cleavage furrows  where small particles fuse into large ones. The loss of maternal rbm24a causes the complete degradation of germ plasm components and the disappearance of PGCs  resulting in totally sterile animals. Our work establishes that Rbm24 is a critical nucleating organizer component of germ plasm  highlighting an emerging common mechanism to read and recruit RNA component into phase separated condensates. Overall design: To comprehensively characterize the RNA binding landscape of Rbm24a in zebrafish  we performed RIP seq analysis using rbm24a GFP knock in samples  with IgG serving as the control.,,,,Zebrafish  RIP seq  Rbm24a Knock in2 IgG2,GSM8759067,,source name:whole embryo|tissue:whole embryo|cell line:4 cell stage|cell type:embryonic cell|genotype:knock in|geo loc name:missing|collection date:missing,Zebrafish  RIP seq  Rbm24a Knock in2 IgG2,Each library was amplified by a 15 cycle PCR  and 150 bp paired end sequencing was subjected to Illumina Nova seq 6000 to obtain the raw data. Assembly: GRCz11 Supplementary files format and content: tab delimited text file includes raw counts for each Sample Supplementary files format and content: Bigwig file for each Sample,whole embryo,,Total RNAs were extracted from Mrbm24a embryos at 4 cell sphere 24 hpf stage  using TRIzol reagent Invitrogen  and were precipitated by cold isopropanol 50%  v/v in the presence of carrier glycogen 20 µg/each sample. 1 μg of immunoprecipitated RNA was utilized for library construction. Ribosomal RNA depletion was performed using RNase H based probes followed by magnetic bead purification. RNA fragmentation was achieved through magnesium catalyzed hydrolysis under elevated temperature 94°C for 6 min. First strand cDNA synthesis was initiated with random hexamer primers and reverse transcriptase  followed by second strand synthesis using DNA polymerase I with dUTP incorporation for strand marking. Double stranded cDNA fragments underwent end repair and three prime adenylation using a single tube enzymatic reaction system. Illumina compatible adaptors with unique dual indexes were ligated to both ends of the DNA fragments. Size selection of adaptor ligated products 180 250 bp was performed using double sided SPRI bead purification. PCR amplification was conducted with P5/P7 primers for 12 cycles  followed by AMPure XP bead purification to remove primer dimers. Library quality was verified by Agilent 2100 Bioanalyzer DNA High Sensitivity chip analysis. Indexed libraries were pooled in equimolar ratios and subjected to paired end sequencing 2×150 bp on an Illumina NovaSeq 6000 platform following the manufacturer's standardized workflow.,Zebrafish embryos were maintained in 1/3x Ringer's at 28.5C,tissue:whole embryo|cell line:4 cell stage|cell type:embryonic cell|genotype:knock in,GSM8759067,GSM8759067: Zebrafish  RIP seq  Rbm24a Knock in2 IgG2; Danio rerio; RIP Seq,GSM8759067 r1,GSM8759067,1,Total RNAs were extracted from Mrbm24a embryos at 4 cell sphere 24 hpf stage  using TRIzol reagent Invitrogen  and were precipitated by cold isopropanol 50%  v/v in the presence of carrier glycogen 20 µg/each sample. 1 μg of immunoprecipitated RNA was utilized for library construction. Ribosomal RNA depletion was performed using RNase H based probes followed by magnetic bead purification. RNA fragmentation was achieved through magnesium catalyzed hydrolysis under elevated temperature 94°C for 6 min. First strand cDNA synthesis was initiated with random hexamer primers and reverse transcriptase  followed by second strand synthesis using DNA polymerase I with dUTP incorporation for strand marking. Double stranded cDNA fragments underwent end repair and three prime adenylation using a single tube enzymatic reaction system. Illumina compatible adaptors with unique dual indexes were ligated to both ends of the DNA fragments. Size selection of adaptor ligated products 180 250 bp was performed using double sided SPRI bead purification. PCR amplification was conducted with P5/P7 primers for 12 cycles  followed by AMPure XP bead purification to remove primer dimers. Library quality was verified by Agilent 2100 Bioanalyzer DNA High Sensitivity chip analysis. Indexed libraries were pooled in equimolar ratios and subjected to paired end sequencing 2×150 bp on an Illumina NovaSeq 6000 platform following the manufacturer's standardized workflow.,,RIP-Seq,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP560121,,,RIP_IgG2.R1.fq.gz RIP_IgG2.R2.fq.gz,fastq fastq,18588838088.0,61552444.0,GSM8759067 r1,0:151 1:151,A:3864885070;C:5250483318;G:5713729791;T:3756127723;N:3612186,151,151,,,3864885070,5250483318,5713729791,3756127723,3612186,SRX27483375,SRS23906279,SRA2061743,"Ang Li, College of Life Sciences, shandong university","Ang Li, College of Life Sciences, shandong university",,,,,,,,,,,,B,T,mate2 technical by mapping diff,illumina,novaseq_era,unknown,random_priming,unknown,bulk,unknown,unknown,,China,2025-01-27,Multi-stage,Embryo,Whole Organism,All anatomical structures
34617,SRR32137216,SRX27483374,SRS23906278,SRP560121,PRJNA1216205,rbm24a is an organizer component of germ plasm to determine germ cell fate 202409 validation,GSE288109,Other,The  formation of germ cells is a critical issue for the continuation of species. A large group of animals follow a preformation strategy to generate their primordial germ cells PGCs. They produce a set of localized maternal mRNAs and proteins to form phase separated germ plasm that functions as the PGC determinants  but the mechanisms underlying the assembly of germ plasm is poorly understood. This study identifies Rbm24a as an enssential localized germ plasm protein component that controls the formation of large and functional germ plasm granules. Rbm24a is complexed with Buc and interacts with germ plasm mRNAs  which determines the specific grasp of germ plasm mRNAs into the phase separated aggregates. Rbm24a absent granules fail to undergo kinesin dependent transport towards the cleavage furrows  where small particles fuse into large ones. The loss of maternal rbm24a causes the complete degradation of germ plasm components and the disappearance of PGCs  resulting in totally sterile animals. Our work establishes that Rbm24 is a critical nucleating organizer component of germ plasm  highlighting an emerging common mechanism to read and recruit RNA component into phase separated condensates. Overall design: To comprehensively characterize the RNA binding landscape of Rbm24a in zebrafish  we performed RIP seq analysis using rbm24a GFP knock in samples  with IgG serving as the control.,,,,Zebrafish  RIP seq  Rbm24a Knock in1 IgG1,GSM8759066,,source name:whole embryo|tissue:whole embryo|cell line:4 cell stage|cell type:embryonic cell|genotype:knock in|geo loc name:missing|collection date:missing,Zebrafish  RIP seq  Rbm24a Knock in1 IgG1,Each library was amplified by a 15 cycle PCR  and 150 bp paired end sequencing was subjected to Illumina Nova seq 6000 to obtain the raw data. Assembly: GRCz11 Supplementary files format and content: tab delimited text file includes raw counts for each Sample Supplementary files format and content: Bigwig file for each Sample,whole embryo,,Total RNAs were extracted from Mrbm24a embryos at 4 cell sphere 24 hpf stage  using TRIzol reagent Invitrogen  and were precipitated by cold isopropanol 50%  v/v in the presence of carrier glycogen 20 µg/each sample. 1 μg of immunoprecipitated RNA was utilized for library construction. Ribosomal RNA depletion was performed using RNase H based probes followed by magnetic bead purification. RNA fragmentation was achieved through magnesium catalyzed hydrolysis under elevated temperature 94°C for 6 min. First strand cDNA synthesis was initiated with random hexamer primers and reverse transcriptase  followed by second strand synthesis using DNA polymerase I with dUTP incorporation for strand marking. Double stranded cDNA fragments underwent end repair and three prime adenylation using a single tube enzymatic reaction system. Illumina compatible adaptors with unique dual indexes were ligated to both ends of the DNA fragments. Size selection of adaptor ligated products 180 250 bp was performed using double sided SPRI bead purification. PCR amplification was conducted with P5/P7 primers for 12 cycles  followed by AMPure XP bead purification to remove primer dimers. Library quality was verified by Agilent 2100 Bioanalyzer DNA High Sensitivity chip analysis. Indexed libraries were pooled in equimolar ratios and subjected to paired end sequencing 2×150 bp on an Illumina NovaSeq 6000 platform following the manufacturer's standardized workflow.,Zebrafish embryos were maintained in 1/3x Ringer's at 28.5C,tissue:whole embryo|cell line:4 cell stage|cell type:embryonic cell|genotype:knock in,GSM8759066,GSM8759066: Zebrafish  RIP seq  Rbm24a Knock in1 IgG1; Danio rerio; RIP Seq,GSM8759066 r1,GSM8759066,1,Total RNAs were extracted from Mrbm24a embryos at 4 cell sphere 24 hpf stage  using TRIzol reagent Invitrogen  and were precipitated by cold isopropanol 50%  v/v in the presence of carrier glycogen 20 µg/each sample. 1 μg of immunoprecipitated RNA was utilized for library construction. Ribosomal RNA depletion was performed using RNase H based probes followed by magnetic bead purification. RNA fragmentation was achieved through magnesium catalyzed hydrolysis under elevated temperature 94°C for 6 min. First strand cDNA synthesis was initiated with random hexamer primers and reverse transcriptase  followed by second strand synthesis using DNA polymerase I with dUTP incorporation for strand marking. Double stranded cDNA fragments underwent end repair and three prime adenylation using a single tube enzymatic reaction system. Illumina compatible adaptors with unique dual indexes were ligated to both ends of the DNA fragments. Size selection of adaptor ligated products 180 250 bp was performed using double sided SPRI bead purification. PCR amplification was conducted with P5/P7 primers for 12 cycles  followed by AMPure XP bead purification to remove primer dimers. Library quality was verified by Agilent 2100 Bioanalyzer DNA High Sensitivity chip analysis. Indexed libraries were pooled in equimolar ratios and subjected to paired end sequencing 2×150 bp on an Illumina NovaSeq 6000 platform following the manufacturer's standardized workflow.,,RIP-Seq,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP560121,,,RIP_IgG1.R1.fq.gz RIP_IgG1.R2.fq.gz,fastq fastq,18008086350.0,59629425.0,GSM8759066 r1,0:151 1:151,A:3736851298;C:5091099145;G:5529269310;T:3647429389;N:3437208,151,151,,,3736851298,5091099145,5529269310,3647429389,3437208,SRX27483374,SRS23906278,SRA2061743,"Ang Li, College of Life Sciences, shandong university","Ang Li, College of Life Sciences, shandong university",,,,,,,,,,,,B,T,mate2 technical by mapping diff,illumina,novaseq_era,unknown,random_priming,unknown,bulk,unknown,unknown,,China,2025-01-27,Multi-stage,Embryo,Whole Organism,All anatomical structures
36053,SRR33444765,SRX28685529,SRS24952431,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 8 dimethylnaphthalene  A,GSM8969898,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 8 dimethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,1 8 dimethylnaphthalene  A,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 8 dimethylnaphthalene|batch:1,GSM8969898,GSM8969898: 1 8 dimethylnaphthalene  A; Danio rerio; OTHER,GSM8969898 r1,GSM8969898,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,2_A.fastq.gz,fastq,610412472.0,11968872.0,GSM8969898 r1,0:51,A:127080561;C:159924680;G:139627579;T:183641966;N:137686,51,,,,127080561,159924680,139627579,183641966,137686,SRX28685529,SRS24952431,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36054,SRR33444766,SRX28685528,SRS24952432,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,control day 2  C,GSM8969935,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:control day 2|batch:2|geo loc name:missing|collection date:missing,control day 2  C,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:control day 2|batch:2,GSM8969935,GSM8969935: control day 2  C; Danio rerio; OTHER,GSM8969935 r1,GSM8969935,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,C2C.fastq.gz,fastq,741413877.0,14537527.0,GSM8969935 r1,0:51,A:153346994;C:193370788;G:172893997;T:221634171;N:167927,51,,,,153346994,193370788,172893997,221634171,167927,SRX28685528,SRS24952432,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36055,SRR33444767,SRX28685527,SRS24952430,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,control day 2  B,GSM8969934,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:control day 2|batch:2|geo loc name:missing|collection date:missing,control day 2  B,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:control day 2|batch:2,GSM8969934,GSM8969934: control day 2  B; Danio rerio; OTHER,GSM8969934 r1,GSM8969934,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,C2B.fastq.gz,fastq,541591338.0,10619438.0,GSM8969934 r1,0:51,A:110558693;C:142536702;G:127264949;T:161109133;N:121861,51,,,,110558693,142536702,127264949,161109133,121861,SRX28685527,SRS24952430,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36056,SRR33444768,SRX28685526,SRS24952429,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,control day 2  A,GSM8969933,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:control day 2|batch:2|geo loc name:missing|collection date:missing,control day 2  A,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:control day 2|batch:2,GSM8969933,GSM8969933: control day 2  A; Danio rerio; OTHER,GSM8969933 r1,GSM8969933,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,C2A.fastq.gz,fastq,613454112.0,12028512.0,GSM8969933 r1,0:51,A:125133985;C:160879875;G:143614243;T:183687282;N:138727,51,,,,125133985,160879875,143614243,183687282,138727,SRX28685526,SRS24952429,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36057,SRR33444769,SRX28685525,SRS24952428,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,control day 1  C,GSM8969932,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:control day 1|batch:1|geo loc name:missing|collection date:missing,control day 1  C,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:control day 1|batch:1,GSM8969932,GSM8969932: control day 1  C; Danio rerio; OTHER,GSM8969932 r1,GSM8969932,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,C1C.fastq.gz,fastq,474767415.0,9309165.0,GSM8969932 r1,0:51,A:97071261;C:124093451;G:110241992;T:143253948;N:106763,51,,,,97071261,124093451,110241992,143253948,106763,SRX28685525,SRS24952428,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36058,SRR33444770,SRX28685524,SRS24952427,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,control day 1  B,GSM8969931,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:control day 1|batch:1|geo loc name:missing|collection date:missing,control day 1  B,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:control day 1|batch:1,GSM8969931,GSM8969931: control day 1  B; Danio rerio; OTHER,GSM8969931 r1,GSM8969931,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,C1B.fastq.gz,fastq,457427772.0,8969172.0,GSM8969931 r1,0:51,A:94621194;C:123105730;G:103779476;T:135818957;N:102415,51,,,,94621194,123105730,103779476,135818957,102415,SRX28685524,SRS24952427,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36059,SRR33444771,SRX28685523,SRS24952426,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,control day 1  A,GSM8969930,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:control day 1|batch:1|geo loc name:missing|collection date:missing,control day 1  A,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:control day 1|batch:1,GSM8969930,GSM8969930: control day 1  A; Danio rerio; OTHER,GSM8969930 r1,GSM8969930,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,C1A.fastq.gz,fastq,492735021.0,9661471.0,GSM8969930 r1,0:51,A:101186692;C:127371147;G:115048213;T:149017307;N:111662,51,,,,101186692,127371147,115048213,149017307,111662,SRX28685523,SRS24952426,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36060,SRR33444772,SRX28685522,SRS24952423,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 6 7 trimethylnaphthalene  E,GSM8969929,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 6 7 trimethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,1 6 7 trimethylnaphthalene  E,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 6 7 trimethylnaphthalene|batch:1,GSM8969929,GSM8969929: 1 6 7 trimethylnaphthalene  E; Danio rerio; OTHER,GSM8969929 r1,GSM8969929,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,9_E.fastq.gz,fastq,611528913.0,11990763.0,GSM8969929 r1,0:51,A:124350552;C:160747373;G:139974969;T:186316449;N:139570,51,,,,124350552,160747373,139974969,186316449,139570,SRX28685522,SRS24952423,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36061,SRR33444773,SRX28685521,SRS24952425,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 6 7 trimethylnaphthalene  D,GSM8969928,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 6 7 trimethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,1 6 7 trimethylnaphthalene  D,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 6 7 trimethylnaphthalene|batch:1,GSM8969928,GSM8969928: 1 6 7 trimethylnaphthalene  D; Danio rerio; OTHER,GSM8969928 r1,GSM8969928,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,9_D.fastq.gz,fastq,628858662.0,12330562.0,GSM8969928 r1,0:51,A:128538964;C:164901356;G:144434115;T:190840964;N:143263,51,,,,128538964,164901356,144434115,190840964,143263,SRX28685521,SRS24952425,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36062,SRR33444774,SRX28685520,SRS24952424,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 6 7 trimethylnaphthalene  B,GSM8969927,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 6 7 trimethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,1 6 7 trimethylnaphthalene  B,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 6 7 trimethylnaphthalene|batch:1,GSM8969927,GSM8969927: 1 6 7 trimethylnaphthalene  B; Danio rerio; OTHER,GSM8969927 r1,GSM8969927,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,9_B.fastq.gz,fastq,550098036.0,10786236.0,GSM8969927 r1,0:51,A:112188300;C:144676062;G:126419391;T:166689819;N:124464,51,,,,112188300,144676062,126419391,166689819,124464,SRX28685520,SRS24952424,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36063,SRR33444775,SRX28685519,SRS24952422,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 6 7 trimethylnaphthalene  A,GSM8969926,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 6 7 trimethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,1 6 7 trimethylnaphthalene  A,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 6 7 trimethylnaphthalene|batch:1,GSM8969926,GSM8969926: 1 6 7 trimethylnaphthalene  A; Danio rerio; OTHER,GSM8969926 r1,GSM8969926,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,9_A.fastq.gz,fastq,449907771.0,8821721.0,GSM8969926 r1,0:51,A:91463949;C:117779693;G:103518447;T:137043427;N:102255,51,,,,91463949,117779693,103518447,137043427,102255,SRX28685519,SRS24952422,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36064,SRR33444776,SRX28685518,SRS24952421,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 ethylnaphthalene  E,GSM8969925,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 ethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,1 ethylnaphthalene  E,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 ethylnaphthalene|batch:1,GSM8969925,GSM8969925: 1 ethylnaphthalene  E; Danio rerio; OTHER,GSM8969925 r1,GSM8969925,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,8_E.fastq.gz,fastq,596647062.0,11698962.0,GSM8969925 r1,0:51,A:121970662;C:156141284;G:138301387;T:180098649;N:135080,51,,,,121970662,156141284,138301387,180098649,135080,SRX28685518,SRS24952421,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36065,SRR33444777,SRX28685517,SRS24952420,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 ethylnaphthalene  C,GSM8969924,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 ethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,1 ethylnaphthalene  C,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 ethylnaphthalene|batch:1,GSM8969924,GSM8969924: 1 ethylnaphthalene  C; Danio rerio; OTHER,GSM8969924 r1,GSM8969924,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,8_C.fastq.gz,fastq,659636703.0,12934053.0,GSM8969924 r1,0:51,A:134295377;C:173327337;G:153006654;T:198857895;N:149440,51,,,,134295377,173327337,153006654,198857895,149440,SRX28685517,SRS24952420,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36066,SRR33444778,SRX28685516,SRS24952418,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 ethylnaphthalene  B,GSM8969923,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 ethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,1 ethylnaphthalene  B,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 ethylnaphthalene|batch:1,GSM8969923,GSM8969923: 1 ethylnaphthalene  B; Danio rerio; OTHER,GSM8969923 r1,GSM8969923,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,8_B.fastq.gz,fastq,748961214.0,14685514.0,GSM8969923 r1,0:51,A:153969741;C:195524281;G:173004091;T:226292712;N:170389,51,,,,153969741,195524281,173004091,226292712,170389,SRX28685516,SRS24952418,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36067,SRR33444779,SRX28685515,SRS24952419,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 ethylnaphthalene  A,GSM8969922,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 ethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,1 ethylnaphthalene  A,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 ethylnaphthalene|batch:1,GSM8969922,GSM8969922: 1 ethylnaphthalene  A; Danio rerio; OTHER,GSM8969922 r1,GSM8969922,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,8_A.fastq.gz,fastq,587085225.0,11511475.0,GSM8969922 r1,0:51,A:119552515;C:153868067;G:135452522;T:178079346;N:132775,51,,,,119552515,153868067,135452522,178079346,132775,SRX28685515,SRS24952419,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36068,SRR33444780,SRX28685514,SRS24952417,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,naphthalene  D,GSM8969889,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:naphthalene|batch:2|geo loc name:missing|collection date:missing,naphthalene  D,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:naphthalene|batch:2,GSM8969889,GSM8969889: naphthalene  D; Danio rerio; OTHER,GSM8969889 r1,GSM8969889,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,20D.fastq.gz,fastq,574442223.0,11263573.0,GSM8969889 r1,0:51,A:115679820;C:152904111;G:134156914;T:171570534;N:130844,51,,,,115679820,152904111,134156914,171570534,130844,SRX28685514,SRS24952417,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36069,SRR33444781,SRX28685513,SRS24952416,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,naphthalene  C,GSM8969888,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:naphthalene|batch:2|geo loc name:missing|collection date:missing,naphthalene  C,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:naphthalene|batch:2,GSM8969888,GSM8969888: naphthalene  C; Danio rerio; OTHER,GSM8969888 r1,GSM8969888,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,20C.fastq.gz,fastq,626063199.0,12275749.0,GSM8969888 r1,0:51,A:126797180;C:165522830;G:146080993;T:187520712;N:141484,51,,,,126797180,165522830,146080993,187520712,141484,SRX28685513,SRS24952416,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36070,SRR33444782,SRX28685512,SRS24952415,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,naphthalene  B,GSM8969887,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:naphthalene|batch:2|geo loc name:missing|collection date:missing,naphthalene  B,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:naphthalene|batch:2,GSM8969887,GSM8969887: naphthalene  B; Danio rerio; OTHER,GSM8969887 r1,GSM8969887,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,20B.fastq.gz,fastq,743186535.0,14572285.0,GSM8969887 r1,0:51,A:149056375;C:196734696;G:173272512;T:223954929;N:168023,51,,,,149056375,196734696,173272512,223954929,168023,SRX28685512,SRS24952415,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36071,SRR33444783,SRX28685511,SRS24952413,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,naphthalene  A,GSM8969886,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:naphthalene|batch:2|geo loc name:missing|collection date:missing,naphthalene  A,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:naphthalene|batch:2,GSM8969886,GSM8969886: naphthalene  A; Danio rerio; OTHER,GSM8969886 r1,GSM8969886,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,20A.fastq.gz,fastq,586121376.0,11492576.0,GSM8969886 r1,0:51,A:117510651;C:155746473;G:136839801;T:175891905;N:132546,51,,,,117510651,155746473,136839801,175891905,132546,SRX28685511,SRS24952413,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36072,SRR33444784,SRX28685510,SRS24952414,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,2 6 diethylnaphthalene  E,GSM8969885,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:2 6 diethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,2 6 diethylnaphthalene  E,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:2 6 diethylnaphthalene|batch:1,GSM8969885,GSM8969885: 2 6 diethylnaphthalene  E; Danio rerio; OTHER,GSM8969885 r1,GSM8969885,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,1_E.fastq.gz,fastq,554540646.0,10873346.0,GSM8969885 r1,0:51,A:113384528;C:146196077;G:126957362;T:167877836;N:124843,51,,,,113384528,146196077,126957362,167877836,124843,SRX28685510,SRS24952414,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36073,SRR33444785,SRX28685509,SRS24952412,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,2 6 diethylnaphthalene  1D,GSM8969884,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:2 6 diethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,2 6 diethylnaphthalene  1D,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:2 6 diethylnaphthalene|batch:1,GSM8969884,GSM8969884: 2 6 diethylnaphthalene  1D; Danio rerio; OTHER,GSM8969884 r1,GSM8969884,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,1_D.fastq.gz,fastq,663557685.0,13010935.0,GSM8969884 r1,0:51,A:137450973;C:174284217;G:151682321;T:199990487;N:149687,51,,,,137450973,174284217,151682321,199990487,149687,SRX28685509,SRS24952412,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36074,SRR33444786,SRX28685508,SRS24952410,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,2 6 diethylnaphthalene  C,GSM8969883,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:2 6 diethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,2 6 diethylnaphthalene  C,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:2 6 diethylnaphthalene|batch:1,GSM8969883,GSM8969883: 2 6 diethylnaphthalene  C; Danio rerio; OTHER,GSM8969883 r1,GSM8969883,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,1_C.fastq.gz,fastq,571682562.0,11209462.0,GSM8969883 r1,0:51,A:118075588;C:149868301;G:131235979;T:172374281;N:128413,51,,,,118075588,149868301,131235979,172374281,128413,SRX28685508,SRS24952410,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36075,SRR33444787,SRX28685507,SRS24952411,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,2 6 diethylnaphthalene  B,GSM8969882,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:2 6 diethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,2 6 diethylnaphthalene  B,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:2 6 diethylnaphthalene|batch:1,GSM8969882,GSM8969882: 2 6 diethylnaphthalene  B; Danio rerio; OTHER,GSM8969882 r1,GSM8969882,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,1_B.fastq.gz,fastq,431385387.0,8458537.0,GSM8969882 r1,0:51,A:89066718;C:112472814;G:98920613;T:130827111;N:98131,51,,,,89066718,112472814,98920613,130827111,98131,SRX28685507,SRS24952411,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36076,SRR33444788,SRX28685506,SRS24952409,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,2 4 5 trimethylnaphthalene  E,GSM8969921,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:2 4 5 trimethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,2 4 5 trimethylnaphthalene  E,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:2 4 5 trimethylnaphthalene|batch:1,GSM8969921,GSM8969921: 2 4 5 trimethylnaphthalene  E; Danio rerio; OTHER,GSM8969921 r1,GSM8969921,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,7_E.fastq.gz,fastq,674676552.0,13228952.0,GSM8969921 r1,0:51,A:137983580;C:176549665;G:153689206;T:206301698;N:152403,51,,,,137983580,176549665,153689206,206301698,152403,SRX28685506,SRS24952409,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36077,SRR33444789,SRX28685505,SRS24952408,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,2 4 5 trimethylnaphthalene  D,GSM8969920,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:2 4 5 trimethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,2 4 5 trimethylnaphthalene  D,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:2 4 5 trimethylnaphthalene|batch:1,GSM8969920,GSM8969920: 2 4 5 trimethylnaphthalene  D; Danio rerio; OTHER,GSM8969920 r1,GSM8969920,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,7_D.fastq.gz,fastq,585836439.0,11486989.0,GSM8969920 r1,0:51,A:119799324;C:153323440;G:134779037;T:177801818;N:132820,51,,,,119799324,153323440,134779037,177801818,132820,SRX28685505,SRS24952408,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36078,SRR33444790,SRX28685504,SRS24952406,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,2 4 5 trimethylnaphthalene  C,GSM8969919,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:2 4 5 trimethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,2 4 5 trimethylnaphthalene  C,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:2 4 5 trimethylnaphthalene|batch:1,GSM8969919,GSM8969919: 2 4 5 trimethylnaphthalene  C; Danio rerio; OTHER,GSM8969919 r1,GSM8969919,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,7_C.fastq.gz,fastq,528946500.0,10371500.0,GSM8969919 r1,0:51,A:108461005;C:138556942;G:121948616;T:159859746;N:120191,51,,,,108461005,138556942,121948616,159859746,120191,SRX28685504,SRS24952406,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36079,SRR33444791,SRX28685503,SRS24952407,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,2 4 5 trimethylnaphthalene  B,GSM8969918,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:2 4 5 trimethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,2 4 5 trimethylnaphthalene  B,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:2 4 5 trimethylnaphthalene|batch:1,GSM8969918,GSM8969918: 2 4 5 trimethylnaphthalene  B; Danio rerio; OTHER,GSM8969918 r1,GSM8969918,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,7_B.fastq.gz,fastq,477634431.0,9365381.0,GSM8969918 r1,0:51,A:96765678;C:126003340;G:110639151;T:144117143;N:109119,51,,,,96765678,126003340,110639151,144117143,109119,SRX28685503,SRS24952407,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36080,SRR33444792,SRX28685502,SRS24952405,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 4 5 trimethylnaphthalene  E,GSM8969917,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 4 5 trimethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,1 4 5 trimethylnaphthalene  E,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 4 5 trimethylnaphthalene|batch:1,GSM8969917,GSM8969917: 1 4 5 trimethylnaphthalene  E; Danio rerio; OTHER,GSM8969917 r1,GSM8969917,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,6_E.fastq.gz,fastq,611799264.0,11996064.0,GSM8969917 r1,0:51,A:124227226;C:161277466;G:140911377;T:185244748;N:138447,51,,,,124227226,161277466,140911377,185244748,138447,SRX28685502,SRS24952405,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36081,SRR33444793,SRX28685501,SRS24952404,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 4 5 trimethylnaphthalene  D,GSM8969916,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 4 5 trimethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,1 4 5 trimethylnaphthalene  D,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 4 5 trimethylnaphthalene|batch:1,GSM8969916,GSM8969916: 1 4 5 trimethylnaphthalene  D; Danio rerio; OTHER,GSM8969916 r1,GSM8969916,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,6_D.fastq.gz,fastq,641235036.0,12573236.0,GSM8969916 r1,0:51,A:131787611;C:167389654;G:147724133;T:194188316;N:145322,51,,,,131787611,167389654,147724133,194188316,145322,SRX28685501,SRS24952404,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36082,SRR33444794,SRX28685500,SRS24952403,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 4 5 trimethylnaphthalene  C,GSM8969915,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 4 5 trimethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,1 4 5 trimethylnaphthalene  C,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 4 5 trimethylnaphthalene|batch:1,GSM8969915,GSM8969915: 1 4 5 trimethylnaphthalene  C; Danio rerio; OTHER,GSM8969915 r1,GSM8969915,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,6_C.fastq.gz,fastq,729657561.0,14307011.0,GSM8969915 r1,0:51,A:149650263;C:189405599;G:167732880;T:222702499;N:166320,51,,,,149650263,189405599,167732880,222702499,166320,SRX28685500,SRS24952403,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36083,SRR33444795,SRX28685499,SRS24952402,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 4 5 trimethylnaphthalene  A,GSM8969914,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 4 5 trimethylnaphthalene|batch:1|geo loc name:missing|collection date:missing,1 4 5 trimethylnaphthalene  A,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 4 5 trimethylnaphthalene|batch:1,GSM8969914,GSM8969914: 1 4 5 trimethylnaphthalene  A; Danio rerio; OTHER,GSM8969914 r1,GSM8969914,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,6_A.fastq.gz,fastq,605891169.0,11880219.0,GSM8969914 r1,0:51,A:124734415;C:158082271;G:139532340;T:183405053;N:137090,51,,,,124734415,158082271,139532340,183405053,137090,SRX28685499,SRS24952402,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36084,SRR33444796,SRX28685498,SRS24952401,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,2 butylnaphthalene  E,GSM8969881,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:2 butylnaphthalene|batch:2|geo loc name:missing|collection date:missing,2 butylnaphthalene  E,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:2 butylnaphthalene|batch:2,GSM8969881,GSM8969881: 2 butylnaphthalene  E; Danio rerio; OTHER,GSM8969881 r1,GSM8969881,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,19E.fastq.gz,fastq,680434299.0,13341849.0,GSM8969881 r1,0:51,A:137122133;C:179581944;G:158458965;T:205117134;N:154123,51,,,,137122133,179581944,158458965,205117134,154123,SRX28685498,SRS24952401,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36085,SRR33444797,SRX28685497,SRS24952400,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,2 butylnaphthalene  C,GSM8969880,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:2 butylnaphthalene|batch:2|geo loc name:missing|collection date:missing,2 butylnaphthalene  C,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:2 butylnaphthalene|batch:2,GSM8969880,GSM8969880: 2 butylnaphthalene  C; Danio rerio; OTHER,GSM8969880 r1,GSM8969880,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,19C.fastq.gz,fastq,550652814.0,10797114.0,GSM8969880 r1,0:51,A:110713427;C:146318389;G:128285092;T:165211419;N:124487,51,,,,110713427,146318389,128285092,165211419,124487,SRX28685497,SRS24952400,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36086,SRR33444798,SRX28685496,SRS24952399,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,2 butylnaphthalene  B,GSM8969879,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:2 butylnaphthalene|batch:2|geo loc name:missing|collection date:missing,2 butylnaphthalene  B,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:2 butylnaphthalene|batch:2,GSM8969879,GSM8969879: 2 butylnaphthalene  B; Danio rerio; OTHER,GSM8969879 r1,GSM8969879,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,19B.fastq.gz,fastq,534179763.0,10474113.0,GSM8969879 r1,0:51,A:107605335;C:141768628;G:124858929;T:159826026;N:120845,51,,,,107605335,141768628,124858929,159826026,120845,SRX28685496,SRS24952399,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36087,SRR33444799,SRX28685495,SRS24952398,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,2 butylnaphthalene  A,GSM8969878,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:2 butylnaphthalene|batch:2|geo loc name:missing|collection date:missing,2 butylnaphthalene  A,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:2 butylnaphthalene|batch:2,GSM8969878,GSM8969878: 2 butylnaphthalene  A; Danio rerio; OTHER,GSM8969878 r1,GSM8969878,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,19A.fastq.gz,fastq,512341716.0,10045916.0,GSM8969878 r1,0:51,A:102951861;C:135598511;G:120056781;T:153618339;N:116224,51,,,,102951861,135598511,120056781,153618339,116224,SRX28685495,SRS24952398,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36088,SRR33444800,SRX28685494,SRS24952396,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 6 dimethylnaphthalene  E,GSM8969877,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 6 dimethylnaphthalene|batch:2|geo loc name:missing|collection date:missing,1 6 dimethylnaphthalene  E,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 6 dimethylnaphthalene|batch:2,GSM8969877,GSM8969877: 1 6 dimethylnaphthalene  E; Danio rerio; OTHER,GSM8969877 r1,GSM8969877,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,18E.fastq.gz,fastq,533747334.0,10465634.0,GSM8969877 r1,0:51,A:106676262;C:142402611;G:124929407;T:159618343;N:120711,51,,,,106676262,142402611,124929407,159618343,120711,SRX28685494,SRS24952396,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36089,SRR33444801,SRX28685493,SRS24952397,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 6 dimethylnaphthalene  D,GSM8969876,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 6 dimethylnaphthalene|batch:2|geo loc name:missing|collection date:missing,1 6 dimethylnaphthalene  D,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 6 dimethylnaphthalene|batch:2,GSM8969876,GSM8969876: 1 6 dimethylnaphthalene  D; Danio rerio; OTHER,GSM8969876 r1,GSM8969876,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,18D.fastq.gz,fastq,593617101.0,11639551.0,GSM8969876 r1,0:51,A:120267538;C:157361649;G:138443329;T:177410020;N:134565,51,,,,120267538,157361649,138443329,177410020,134565,SRX28685493,SRS24952397,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36090,SRR33444802,SRX28685492,SRS24952395,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 6 dimethylnaphthalene  C,GSM8969875,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 6 dimethylnaphthalene|batch:2|geo loc name:missing|collection date:missing,1 6 dimethylnaphthalene  C,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 6 dimethylnaphthalene|batch:2,GSM8969875,GSM8969875: 1 6 dimethylnaphthalene  C; Danio rerio; OTHER,GSM8969875 r1,GSM8969875,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,18C.fastq.gz,fastq,699676905.0,13719155.0,GSM8969875 r1,0:51,A:141310126;C:184696569;G:162476370;T:211034854;N:158986,51,,,,141310126,184696569,162476370,211034854,158986,SRX28685492,SRS24952395,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36091,SRR33444803,SRX28685491,SRS24952394,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 6 dimethylnaphthalene  B,GSM8969874,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 6 dimethylnaphthalene|batch:2|geo loc name:missing|collection date:missing,1 6 dimethylnaphthalene  B,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 6 dimethylnaphthalene|batch:2,GSM8969874,GSM8969874: 1 6 dimethylnaphthalene  B; Danio rerio; OTHER,GSM8969874 r1,GSM8969874,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,18B.fastq.gz,fastq,544510578.0,10676678.0,GSM8969874 r1,0:51,A:109680983;C:145022999;G:126931689;T:162751725;N:123182,51,,,,109680983,145022999,126931689,162751725,123182,SRX28685491,SRS24952394,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36092,SRR33444804,SRX28685490,SRS24952393,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 3 dimethylnaphthalene  E,GSM8969865,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 3 dimethylnaphthalene|batch:2|geo loc name:missing|collection date:missing,1 3 dimethylnaphthalene  E,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 3 dimethylnaphthalene|batch:2,GSM8969865,GSM8969865: 1 3 dimethylnaphthalene  E; Danio rerio; OTHER,GSM8969865 r1,GSM8969865,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,15E.fastq.gz,fastq,581996037.0,11411687.0,GSM8969865 r1,0:51,A:115978569;C:155236560;G:135042427;T:175606047;N:132434,51,,,,115978569,155236560,135042427,175606047,132434,SRX28685490,SRS24952393,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36093,SRR33444805,SRX28685489,SRS24952392,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 3 dimethylnaphthalene  D,GSM8969864,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 3 dimethylnaphthalene|batch:2|geo loc name:missing|collection date:missing,1 3 dimethylnaphthalene  D,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 3 dimethylnaphthalene|batch:2,GSM8969864,GSM8969864: 1 3 dimethylnaphthalene  D; Danio rerio; OTHER,GSM8969864 r1,GSM8969864,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,15D.fastq.gz,fastq,586248519.0,11495069.0,GSM8969864 r1,0:51,A:118182908;C:156003232;G:136147171;T:175781962;N:133246,51,,,,118182908,156003232,136147171,175781962,133246,SRX28685489,SRS24952392,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36094,SRR33444806,SRX28685488,SRS24952390,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 3 dimethylnaphthalene  C,GSM8969863,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 3 dimethylnaphthalene|batch:2|geo loc name:missing|collection date:missing,1 3 dimethylnaphthalene  C,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 3 dimethylnaphthalene|batch:2,GSM8969863,GSM8969863: 1 3 dimethylnaphthalene  C; Danio rerio; OTHER,GSM8969863 r1,GSM8969863,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,15C.fastq.gz,fastq,508958835.0,9979585.0,GSM8969863 r1,0:51,A:102290626;C:135909239;G:118336162;T:152307441;N:115367,51,,,,102290626,135909239,118336162,152307441,115367,SRX28685488,SRS24952390,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36095,SRR33444807,SRX28685487,SRS24952391,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,1 3 dimethylnaphthalene  A,GSM8969862,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 3 dimethylnaphthalene|batch:2|geo loc name:missing|collection date:missing,1 3 dimethylnaphthalene  A,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:1 3 dimethylnaphthalene|batch:2,GSM8969862,GSM8969862: 1 3 dimethylnaphthalene  A; Danio rerio; OTHER,GSM8969862 r1,GSM8969862,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,15A.fastq.gz,fastq,486989922.0,9548822.0,GSM8969862 r1,0:51,A:97540748;C:129199875;G:113762642;T:146375255;N:111402,51,,,,97540748,129199875,113762642,146375255,111402,SRX28685487,SRS24952391,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36096,SRR33444808,SRX28685486,SRS24952389,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,2 7 di tert butylnaphthalene  E,GSM8969861,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:2 7 di tert butylnaphthalene|batch:2|geo loc name:missing|collection date:missing,2 7 di tert butylnaphthalene  E,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:2 7 di tert butylnaphthalene|batch:2,GSM8969861,GSM8969861: 2 7 di tert butylnaphthalene  E; Danio rerio; OTHER,GSM8969861 r1,GSM8969861,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,14E.fastq.gz,fastq,585719394.0,11484694.0,GSM8969861 r1,0:51,A:118181050;C:155228152;G:136862691;T:175314585;N:132916,51,,,,118181050,155228152,136862691,175314585,132916,SRX28685486,SRS24952389,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36097,SRR33444809,SRX28685485,SRS24952388,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,2 7 di tert butylnaphthalene  C,GSM8969860,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:2 7 di tert butylnaphthalene|batch:2|geo loc name:missing|collection date:missing,2 7 di tert butylnaphthalene  C,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:2 7 di tert butylnaphthalene|batch:2,GSM8969860,GSM8969860: 2 7 di tert butylnaphthalene  C; Danio rerio; OTHER,GSM8969860 r1,GSM8969860,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,14C.fastq.gz,fastq,668833431.0,13114381.0,GSM8969860 r1,0:51,A:135523737;C:176521149;G:155894870;T:200742014;N:151661,51,,,,135523737,176521149,155894870,200742014,151661,SRX28685485,SRS24952388,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures
36098,SRR33444810,SRX28685484,SRS24952387,SRP583274,PRJNA1259243,Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes,GSE296410,Transcriptome Analysis,Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 µM in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.,,,,2 7 di tert butylnaphthalene  B,GSM8969859,,source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:2 7 di tert butylnaphthalene|batch:2|geo loc name:missing|collection date:missing,2 7 di tert butylnaphthalene  B,Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data,whole animal,When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 µM using DMSO. 10 µL of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28°C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 µL clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 µL RNA shield added. Five pooled samples were collected from each treatment.,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer’s instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,tissue:whole animal|genotype:5D WT|treatment:2 7 di tert butylnaphthalene|batch:2,GSM8969859,GSM8969859: 2 7 di tert butylnaphthalene  B; Danio rerio; OTHER,GSM8969859 r1,GSM8969859,1,1mL of 1µm beads and 500µL of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100µL of lysis buffer was added to tube containing pooled embryo samples and 200µL of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 µL of the supernatant was then transferred to two plates containing 275µL of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50µLof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/µL. 1 µL per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 µL each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.,,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina NovaSeq X,,SRP583274,,,14B.fastq.gz,fastq,745478169.0,14617219.0,GSM8969859 r1,0:51,A:150437358;C:196846497;G:173490170;T:224533891;N:170253,51,,,,150437358,196846497,173490170,224533891,170253,SRX28685484,SRS24952387,SRA2124446,"SARL, Environmental and Molecular Toxicology, Oregon State University","SARL, Environmental and Molecular Toxicology, Oregon State University",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,other,unknown,bulk,unknown,unknown,,United States,2025-05-06,Multi-stage,Embryo,Whole Organism,All anatomical structures