RNAquarium Metadata Server
Institute Website Terms of Use Privacy Policy

run_metadata

151 rows where devstage_curation = "Blastula" and technology = "bulk"

✎ View and edit SQL

This data as json, CSV (advanced)

tissue_curation 2

experiment.platform 2

experiment.library_layout 2

tissue_curation_coarse 1

technology 1

  • bulk · 151 ✖

experiment.library_strategy 1

experiment.library_source 1

experiment.library_selection 1

devstage_curation_coarse 1

devstage_curation 1

  • Blastula · 151 ✖
Link 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
25275 25275 SRR25764093 SRX21486761 SRS18719061 SRP457108 PRJNA1009807 Dynamics of the zebrafish tRNAome during the maternal to zygotic transition [RNA Seq] GSE241752 Transcriptome Analysis 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 RNA seq rep2 GSM7734766   source name:Blastula|strain:TLAB strain|tissue:Blastula|developmental stage:Sphere 4 hpf|genotype:WT|geo loc name:missing|collection date:missing WT sphere 4 hpf RNA seq rep2 3’ adapters were trimmed using Trim Galore v0.6.4 with default settings retaining reads of length ≥20. Reads were aligned to the GRCz11 zebrafish genome using STAR v2.6.1c with the following parameters: outSAMtype BAM SortedByCoordinate outFilterMultimapNmax 1 outFilterMismatchNmax 1 quantMode TranscriptomeSAM GeneCounts alignEndsType Local seedSearchStartLmax 14 alignIntronMax 10000 outFilterIntronMotifs RemoveNoncanonicalUnannotated. featureCounts v1.6.2 was used to count reads overlapping a filtered set of protein coding gene annotations from the GENCODE basic gene annotation. Differential gene expression analysis was performed using DESEq2 v1.38.1 with default settings and gene counts from featureCounts. Assembly: GRCz11 Supplementary files format and content: csv; transcripts per million TPM counts per transcript in MANE annotation Blastula unperturbed growth conditions in E3 medium for zebrafish embryos. 50 whole embryos were collected per sample and immediately lysed in 800uL LiDS/LET buffer5% Lithium dodecyl sulfate in 20 mM Tris HCl pH=7.4 100 mM LiCl 2 mM EDTA 5 mM DTT pH 7.4. 250 ng of the same total RNA used for mim tRNAseq library preparation were used for mRNA Seq library construction with the Zymo Seq RiboFree Total RNA Library Kit Zymo Research #R3000. Libraries were sequenced on a NextSeq 500 platform. 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|genotype:WT GSM7734766 GSM7734766: WT sphere 4 hpf RNA seq rep2; Danio rerio; RNA Seq GSM7734766 r1 GSM7734766 1 50 whole embryos were collected per sample and immediately lysed in 800uL LiDS/LET buffer5% Lithium dodecyl sulfate in 20 mM Tris HCl pH=7.4 100 mM LiCl 2 mM EDTA 5 mM DTT pH 7.4. 250 ng of the same total RNA used for mim tRNAseq library preparation were used for mRNA Seq library construction with the Zymo Seq RiboFree Total RNA Library Kit Zymo Research #R3000. Libraries were sequenced on a NextSeq 500 platform.   RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA NextSeq 500   SRP457108     WT_mRNA_sphere_2.fastq.gz fastq 2455317157.0 30934591.0 GSM7734766 r1 0:79.37 A:614860604;C:588586084;G:522132289;T:729655621;N:82559 79       614860604 588586084 522132289 729655621 82559 SRX21486761 SRS18719061 SRA1700431 Mechanisms of Protein Biogenesis, Max Planck Institute for Biochemistry Max Planck Institute of Biochemistry 1 0.94124   0.10502   0.74876   0.57752   80   B   usable mapping rate illumina nextseq unknown small_rna unknown bulk bulk bulk   Germany 2023-08-28 Blastula Embryo Whole Organism All anatomical structures
25276 25276 SRR25764094 SRX21486760 SRS18719060 SRP457108 PRJNA1009807 Dynamics of the zebrafish tRNAome during the maternal to zygotic transition [RNA Seq] GSE241752 Transcriptome Analysis 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 RNA seq rep1 GSM7734765   source name:Blastula|strain:TLAB strain|tissue:Blastula|developmental stage:Sphere 4 hpf|genotype:WT|geo loc name:missing|collection date:missing WT sphere 4 hpf RNA seq rep1 3’ adapters were trimmed using Trim Galore v0.6.4 with default settings retaining reads of length ≥20. Reads were aligned to the GRCz11 zebrafish genome using STAR v2.6.1c with the following parameters: outSAMtype BAM SortedByCoordinate outFilterMultimapNmax 1 outFilterMismatchNmax 1 quantMode TranscriptomeSAM GeneCounts alignEndsType Local seedSearchStartLmax 14 alignIntronMax 10000 outFilterIntronMotifs RemoveNoncanonicalUnannotated. featureCounts v1.6.2 was used to count reads overlapping a filtered set of protein coding gene annotations from the GENCODE basic gene annotation. Differential gene expression analysis was performed using DESEq2 v1.38.1 with default settings and gene counts from featureCounts. Assembly: GRCz11 Supplementary files format and content: csv; transcripts per million TPM counts per transcript in MANE annotation Blastula unperturbed growth conditions in E3 medium for zebrafish embryos. 50 whole embryos were collected per sample and immediately lysed in 800uL LiDS/LET buffer5% Lithium dodecyl sulfate in 20 mM Tris HCl pH=7.4 100 mM LiCl 2 mM EDTA 5 mM DTT pH 7.4. 250 ng of the same total RNA used for mim tRNAseq library preparation were used for mRNA Seq library construction with the Zymo Seq RiboFree Total RNA Library Kit Zymo Research #R3000. Libraries were sequenced on a NextSeq 500 platform. 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|genotype:WT GSM7734765 GSM7734765: WT sphere 4 hpf RNA seq rep1; Danio rerio; RNA Seq GSM7734765 r1 GSM7734765 1 50 whole embryos were collected per sample and immediately lysed in 800uL LiDS/LET buffer5% Lithium dodecyl sulfate in 20 mM Tris HCl pH=7.4 100 mM LiCl 2 mM EDTA 5 mM DTT pH 7.4. 250 ng of the same total RNA used for mim tRNAseq library preparation were used for mRNA Seq library construction with the Zymo Seq RiboFree Total RNA Library Kit Zymo Research #R3000. Libraries were sequenced on a NextSeq 500 platform.   RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA NextSeq 500   SRP457108     WT_mRNA_sphere_1.fastq.gz fastq 2399640874.0 30232118.0 GSM7734765 r1 0:79.37 A:589069860;C:593745131;G:511439690;T:705305674;N:80519 79       589069860 593745131 511439690 705305674 80519 SRX21486760 SRS18719060 SRA1700431 Mechanisms of Protein Biogenesis, Max Planck Institute for Biochemistry Max Planck Institute of Biochemistry 1 0.85882   0.11848   0.75122   0.5883   80   B   usable mapping rate illumina nextseq unknown small_rna unknown bulk bulk bulk   Germany 2023-08-28 Blastula Embryo Whole Organism All anatomical structures
30050 30050 SRR27730713 SRX23396345 SRS20257259 SRP485064 PRJNA1067370 zebrafish embryo for scRNA seq and bulk RNA seq PRJNA1067370 Other Hepcidin hamp is a key regulator for the maintenance of iron metabolism. How hamp affects the mechanism of zebrafish hematopoiesis is still largely unknown. Here we have generated a stable hamp mutant zebrafish model by using the CRISPR/Cas9 system.We found that iron overload occurred in several tissues of zebrafish and the hemoglobin content decreased significantly during embryonic development.Single cell profiling demonstrated that hematopoietic actors were aberrantly expressed and hematopoietic progenitor cells appeared a group of cell clusters with relatively delayed development accompanied by ferroptosis.       hamp / 4 hpf embryo zebrafish embryo hamp / 4hpf replicate 3   strain:not applicable|isolate:not collected|breed:Danio|cultivar:not applicable|ecotype:not applicable|dev stage:4 hpf|collection date:2022 10|geo loc name:not applicable|sex:not applicable|tissue:embryo|genotype:hamp / |sample type:whole organism|treatment:replicate 3|BioSampleModel:Model organism or animal                 RNA Seq of Danio rerio: 4hpf embryo W 24 W 24 bulk RNA seq     RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina NovaSeq 6000   SRP485064     hamp4h-3_2.fq.gz hamp4h-3_1.fq.gz fastq fastq 6261088500.0 20870295.0 hamp4h 3 1.fq.gz 0:150 1:150 A:1668720393;C:1449635670;G:1495531397;T:1647100279;N:100761 150 150     1668720393 1449635670 1495531397 1647100279 100761 SRX23396345 SRS20257259 SRA1791861 Shanghai Ocean University|College of Fisheries and Life Shanghai Ocean University 2 0.94424 0.94159 0.04146 0.0415 0.75089 0.75201 0.48393 0.48457 150 150 B B biological fallback assumption illumina novaseq_era unknown cdna_unspecified unknown sc_generic bulk bulk   China 2024-01-25 Blastula Embryo Embryo Imprecise All anatomical structures
30061 30061 SRR27730724 SRX23396334 SRS20257248 SRP485064 PRJNA1067370 zebrafish embryo for scRNA seq and bulk RNA seq PRJNA1067370 Other Hepcidin hamp is a key regulator for the maintenance of iron metabolism. How hamp affects the mechanism of zebrafish hematopoiesis is still largely unknown. Here we have generated a stable hamp mutant zebrafish model by using the CRISPR/Cas9 system.We found that iron overload occurred in several tissues of zebrafish and the hemoglobin content decreased significantly during embryonic development.Single cell profiling demonstrated that hematopoietic actors were aberrantly expressed and hematopoietic progenitor cells appeared a group of cell clusters with relatively delayed development accompanied by ferroptosis.       hamp / 4 hpf embryo zebrafish embryo hamp / 4hpf replicate 2   strain:not applicable|isolate:not collected|breed:Danio|cultivar:not applicable|ecotype:not applicable|dev stage:4 hpf|collection date:2022 10|geo loc name:not applicable|sex:not applicable|tissue:embryo|genotype:hamp / |sample type:whole organism|treatment:replicate 2|BioSampleModel:Model organism or animal                 RNA Seq of Danio rerio: 4hpf embryo W 23 W 23 bulk RNA seq     RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina NovaSeq 6000   SRP485064     hamp4h-2_1.fq.gz hamp4h-2_2.fq.gz fastq fastq 6140359800.0 20467866.0 hamp4h 2 1.fq.gz 0:150 1:150 A:1622433757;C:1437175847;G:1481587196;T:1599071088;N:91912 150 150     1622433757 1437175847 1481587196 1599071088 91912 SRX23396334 SRS20257248 SRA1791861 Shanghai Ocean University|College of Fisheries and Life Shanghai Ocean University 2 0.94763 0.94713 0.03975 0.03948 0.75045 0.75193 0.48915 0.48855 150 150 B B biological fallback assumption illumina novaseq_era unknown cdna_unspecified unknown sc_generic bulk bulk   China 2024-01-25 Blastula Embryo Embryo Imprecise All anatomical structures
30062 30062 SRR27730725 SRX23396333 SRS20257247 SRP485064 PRJNA1067370 zebrafish embryo for scRNA seq and bulk RNA seq PRJNA1067370 Other Hepcidin hamp is a key regulator for the maintenance of iron metabolism. How hamp affects the mechanism of zebrafish hematopoiesis is still largely unknown. Here we have generated a stable hamp mutant zebrafish model by using the CRISPR/Cas9 system.We found that iron overload occurred in several tissues of zebrafish and the hemoglobin content decreased significantly during embryonic development.Single cell profiling demonstrated that hematopoietic actors were aberrantly expressed and hematopoietic progenitor cells appeared a group of cell clusters with relatively delayed development accompanied by ferroptosis.       hamp / 4 hpf embryo zebrafish embryo hamp / 4hpf replicate 1   strain:not applicable|isolate:not collected|breed:Danio|cultivar:not applicable|ecotype:not applicable|dev stage:4 hpf|collection date:2022 10|geo loc name:not applicable|sex:not applicable|tissue:embryo|genotype:hamp / |sample type:whole organism|treatment:replicate 1|BioSampleModel:Model organism or animal                 RNA Seq of Danio rerio: 4hpf embryo W 22 W 22 bulk RNA seq     RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina NovaSeq 6000   SRP485064     hamp4h-1_1.fq.gz hamp4h-1_2.fq.gz fastq fastq 6482302500.0 21607675.0 hamp4h 1 1.fq.gz 0:150 1:150 A:1723398342;C:1504238189;G:1551942847;T:1702630056;N:93066 150 150     1723398342 1504238189 1551942847 1702630056 93066 SRX23396333 SRS20257247 SRA1791861 Shanghai Ocean University|College of Fisheries and Life Shanghai Ocean University 2 0.94434 0.94083 0.04297 0.04235 0.75327 0.75526 0.48487 0.4837 150 150 B B biological fallback assumption illumina novaseq_era unknown cdna_unspecified unknown sc_generic bulk bulk   China 2024-01-25 Blastula Embryo Embryo Imprecise All anatomical structures
30063 30063 SRR27674233 SRX23341580 SRS20203706 SRP485064 PRJNA1067370 zebrafish embryo for scRNA seq and bulk RNA seq PRJNA1067370 Other Hepcidin hamp is a key regulator for the maintenance of iron metabolism. How hamp affects the mechanism of zebrafish hematopoiesis is still largely unknown. Here we have generated a stable hamp mutant zebrafish model by using the CRISPR/Cas9 system.We found that iron overload occurred in several tissues of zebrafish and the hemoglobin content decreased significantly during embryonic development.Single cell profiling demonstrated that hematopoietic actors were aberrantly expressed and hematopoietic progenitor cells appeared a group of cell clusters with relatively delayed development accompanied by ferroptosis.       wild type 4 phf embryo zebrafish embryo 4 phf replicate 3   strain:not applicable|isolate:not collected|breed:Danio|cultivar:not applicable|ecotype:not applicable|age:4 phf|dev stage:4 phf|collection date:2022 10|geo loc name:not applicable|sex:not applicable|tissue:embryo|genotype:wild type|sample type:whole organism|treatment:replicate 3|BioSampleModel:Model organism or animal                 RNA Seq of Danio rerio: 4hpf embryo W 3 W 3 bulk RNA seq     RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina NovaSeq 6000   SRP485064     WT4h-3_1.fq.gz WT4h-3_2.fq.gz fastq fastq 5799593100.0 19331977.0 WT4h 3 1.fq.gz 0:150 1:150 A:1543677690;C:1345529657;G:1387996505;T:1522341224;N:48024 150 150     1543677690 1345529657 1387996505 1522341224 48024 SRX23341580 SRS20203706 SRA1789158 Shanghai Ocean University|College of Fisheries and Life Shanghai Ocean University 2 0.94823 0.94693 0.0386 0.03884 0.74686 0.74769 0.47863 0.48203 150 150 B B biological fallback assumption illumina novaseq_era unknown cdna_unspecified unknown sc_generic bulk bulk   China 2024-01-22 Blastula Embryo Embryo Imprecise All anatomical structures
30064 30064 SRR27674234 SRX23341579 SRS20203705 SRP485064 PRJNA1067370 zebrafish embryo for scRNA seq and bulk RNA seq PRJNA1067370 Other Hepcidin hamp is a key regulator for the maintenance of iron metabolism. How hamp affects the mechanism of zebrafish hematopoiesis is still largely unknown. Here we have generated a stable hamp mutant zebrafish model by using the CRISPR/Cas9 system.We found that iron overload occurred in several tissues of zebrafish and the hemoglobin content decreased significantly during embryonic development.Single cell profiling demonstrated that hematopoietic actors were aberrantly expressed and hematopoietic progenitor cells appeared a group of cell clusters with relatively delayed development accompanied by ferroptosis.       wild type 4 phf embryo zebrafish embryo 4 phf replicate 2   strain:not applicable|isolate:not collected|breed:Danio|cultivar:not applicable|ecotype:not applicable|age:4 phf|dev stage:4 phf|collection date:2022 10|geo loc name:not applicable|sex:not applicable|tissue:embryo|genotype:wild type|sample type:whole organism|treatment:replicate 2|BioSampleModel:Model organism or animal                 RNA Seq of Danio rerio: 4hpf embryo W 2 W 2 bulk RNA seq     RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina NovaSeq 6000   SRP485064     WT4h-2_1.fq.gz WT4h-2_2.fq.gz fastq fastq 6408374700.0 21361249.0 WT4h 2 1.fq.gz 0:150 1:150 A:1700326501;C:1489928909;G:1544130015;T:1673936668;N:52607 150 150     1700326501 1489928909 1544130015 1673936668 52607 SRX23341579 SRS20203705 SRA1789158 Shanghai Ocean University|College of Fisheries and Life Shanghai Ocean University 2 0.94833 0.94724 0.03589 0.03528 0.74521 0.74665 0.47991 0.47552 150 150 B B biological fallback assumption illumina novaseq_era unknown cdna_unspecified unknown sc_generic bulk bulk   China 2024-01-22 Blastula Embryo Embryo Imprecise All anatomical structures
30065 30065 SRR27674235 SRX23341578 SRS20203704 SRP485064 PRJNA1067370 zebrafish embryo for scRNA seq and bulk RNA seq PRJNA1067370 Other Hepcidin hamp is a key regulator for the maintenance of iron metabolism. How hamp affects the mechanism of zebrafish hematopoiesis is still largely unknown. Here we have generated a stable hamp mutant zebrafish model by using the CRISPR/Cas9 system.We found that iron overload occurred in several tissues of zebrafish and the hemoglobin content decreased significantly during embryonic development.Single cell profiling demonstrated that hematopoietic actors were aberrantly expressed and hematopoietic progenitor cells appeared a group of cell clusters with relatively delayed development accompanied by ferroptosis.       wild type 4 phf embryo zebrafish embryo 4 phf replicate 1   strain:not applicable|isolate:not collected|breed:Danio|cultivar:not applicable|ecotype:not applicable|age:4 phf|dev stage:4 phf|collection date:2022 10|geo loc name:not applicable|sex:not applicable|tissue:embryo|genotype:wild type|sample type:whole organism|treatment:replicate 1|BioSampleModel:Model organism or animal                 RNA Seq of Danio rerio: 4hpf embryo W 1 W 1 bulk RNA seq     RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina NovaSeq 6000   SRP485064     WT4h-1_1.fq.gz WT4h-1_2.fq.gz fastq fastq 6176335500.0 20587785.0 WT4h 1 1.fq.gz 0:150 1:150 A:1639564255;C:1436792246;G:1481775020;T:1618153065;N:50914 150 150     1639564255 1436792246 1481775020 1618153065 50914 SRX23341578 SRS20203704 SRA1789158 Shanghai Ocean University|College of Fisheries and Life Shanghai Ocean University 2 0.94837 0.94497 0.03675 0.03629 0.74393 0.74497 0.48332 0.48392 150 150 B B biological fallback assumption illumina novaseq_era unknown cdna_unspecified unknown sc_generic bulk bulk   China 2024-01-22 Blastula Embryo Embryo Imprecise All anatomical structures
30749 30749 SRR30167500 SRX25634206 SRS22278604 SRP495323 PRJNA1088158 Protein profiling of zebrafish embryos unmasks regulatory layers during early embryogenesis. GSE261646 Transcriptome Analysis The maternal to zygotic transition is crucial in embryonic development marked by the degradation of maternally provided mRNAs and initiation of zygotic gene expression. However the changes occurring at the protein level during this transition remain unclear. Here we conducted protein profiling throughout zebrafish embryogenesis using quantitative mass spectrometry integrating transcriptomics and translatomics datasets. Our data shows that unlike RNA changes protein changes are less dynamic. Further increases in protein levels correlate with mRNA translation whereas declines in protein levels do not suggesting active protein degradation processes. Interestingly proteins from pure zygotic genes are present at fertilization challenging existing mRNA based gene classifications. As a proof of concept we utilized CRISPR Cas13d to target znf281b mRNA a gene whose protein significantly accumulates within the first two hpf demonstrating its crucial role in development. Consequently our protein profiling coupled with CRISPR Cas13d offers a new approach to unravel maternal mRNAs function during embryonic development. Overall design: Two biological replicate samples containing 50 zebrafish oocytes per stage were collected by pairing females in natural matings to ''purge'' mature eggs and used to establish an oogenesis time line. Fish were euthanized and ovaries harvested within 1–11 days post purging dpp. Stage I and II oocytes were collected at 1–2 dpp stage III oocytes germinal vesicle in central position at 4–7 dpp and stage IV oocytes germinal vesicle asymmetrically located at 8–10 dpp. Oocyte isolations were based on73 and conducted in isolation medium Leibovitz L 15 Sigma Aldrich #L5520 plus Collagenase I and II depending on the stages as follows. Stages I and II were isolated with 3 mg/mL Collagenase I Sigma Aldrich C0130 and 3 mg/mL Collagenase II Gibco 17101015. Stage III was isolated with 3 mg/mL Collagenase I Sigma Aldrich C0130. Stage IV was isolated by mechanical stripping with forceps and nee…   pubmed:39302832   pCas13d al1 4hpf rep3 GSM8446429   source name:Whole embryo|tissue:Whole embryo|developmental stage:4hpf|genotype:AB TF and TLF|geo loc name:missing|collection date:missing pCas13d al1 4hpf rep3 Raw reads from zebrafish oocytes stages I IV and embryos 0 2 4 and 6 hpf were demultiplexed into FASTQ format allowing up to one mismatch using Illumina bcl convert 3.10.5. Reads were aligned to UCSC genome danRer11 with STAR aligner version 2.7.3a using Ensembl 106 gene models. Counts were converted to TPM values using RSEM version v1.3.0 and all subsequent analysis was done using TPM. Assembly: danRer11 Ensembl 106 gene models Supplementary files format and content: comma separated files with TPMs from all replicates Whole embryo   Isolated oocytes were snap frozen and RNA was extracted with TRIzol Invitrogen #15596026 following manufacturer’s protocols. 0 2 4 and 6 hpf zebrafish embryo RNA was extracted with Direct zol RNA MicroPrep kit Zymo Research #R2062 following manufacturer’s protocols. Zebrafish embryos 0 2 4 and 6 hpf total RNA sequencing libraries were generated from 500ng of high quality total RNA as assessed using Bioanalyzer Agilent. Libraries were made according to the manufacturer’s directions for the TruSeq Stranded Total RNA Library Prep Gold Illumina #20020598 and TruSeq RNA Single Indexes Sets A and B Illumina #20020492 and #20020493. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer Agilent Technologies and Qubit Fluorometer Life Technologies. Libraries were pooled quantified and sequenced as 75bp single reads on a high output flow cell using the Illumina NextSeq500 instrument. Following sequencing Illumina Primary Analysis version RTA 2.11.3.0 and bcl convert 3.10.5 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish oocytes stages I IV total RNA Seq libraries were generated from 100 ng of high quality total RNA as assessed by a Bioanalyzer Agilent Technologies. Libraries were prepared according to manufacturer’s instructions using the TruSeq Stranded Total RNA Library Prep Gold Illumina Cat. No. 20020598 and TruSeq RNA Single Indexes; Sets A and B Illumina Cat. No. 20020492 and 20020493. Resulting short fragment libraries were checked for quality and quantity using a Bioanalyzer and a Qubit Fluorometer Life Technologies. Libraries were normalized pooled and sequenced on a NextSeq 500 instrument Illumina as 75 bp single end reads on a high output flowcell using NextSeq Control Software 2.2.0.4. Following sequencing Illumina Primary Analysis version NextSeq RTA 2.4.11 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish znf281b knockdown embryos…   tissue:Whole embryo|developmental stage:4hpf|genotype:AB TF and TLF GSM8446429 GSM8446429: pCas13d al1 4hpf rep3; Danio rerio; RNA Seq GSM8446429 r1 GSM8446429 1 Isolated oocytes were snap frozen and RNA was extracted with TRIzol Invitrogen #15596026 following manufacturer's protocols. 0 2 4 and 6 hpf zebrafish embryo RNA was extracted with Direct zol RNA MicroPrep kit Zymo Research #R2062 following manufacturer's protocols. Zebrafish embryos 0 2 4 and 6 hpf total RNA sequencing libraries were generated from 500ng of high quality total RNA as assessed using Bioanalyzer Agilent. Libraries were made according to the manufacturer's directions for the TruSeq Stranded Total RNA Library Prep Gold Illumina #20020598 and TruSeq RNA Single Indexes Sets A and B Illumina #20020492 and #20020493. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer Agilent Technologies and Qubit Fluorometer Life Technologies. Libraries were pooled quantified and sequenced as 75bp single reads on a high output flow cell using the Illumina NextSeq500 instrument. Following sequencing Illumina Primary Analysis version RTA 2.11.3.0 and bcl convert 3.10.5 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish oocytes stages I IV total RNA Seq libraries were generated from 100 ng of high quality total RNA as assessed by a Bioanalyzer Agilent Technologies. Libraries were prepared according to manufacturer's instructions using the TruSeq Stranded Total RNA Library Prep Gold Illumina Cat. No. 20020598 and TruSeq RNA Single Indexes; Sets A and B Illumina Cat. No. 20020492 and 20020493. Resulting short fragment libraries were checked for quality and quantity using a Bioanalyzer and a Qubit Fluorometer Life Technologies. Libraries were normalized pooled and sequenced on a NextSeq 500 instrument Illumina as 75 bp single end reads on a high output flowcell using NextSeq Control Software 2.2.0.4. Following sequencing Illumina Primary Analysis version NextSeq RTA 2.4.11 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish znf281b knockdown embryos…   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP495323     L67855_S11_L001_R1_001.fastq.gz L67855_S11_L001_R2_001.fastq.gz fastq fastq 2565767500.0 25657675.0 GSM8446429 r1 0:50 1:50 A:676771006;C:596998503;G:603560859;T:688437132;N:0 50 50     676771006 596998503 603560859 688437132 0 SRX25634206 SRS22278604 SRA1942483 Computational Biology, Stowers Institute for Medical Research Computational Biology, Stowers Institute for Medical Research                       B B biological fallback assumption illumina nextseq unknown poly_a trueseq bulk bulk bulk   United States 2024-08-07 Blastula Embryo Whole Organism All anatomical structures
30750 30750 SRR30167501 SRX25634205 SRS22278605 SRP495323 PRJNA1088158 Protein profiling of zebrafish embryos unmasks regulatory layers during early embryogenesis. GSE261646 Transcriptome Analysis The maternal to zygotic transition is crucial in embryonic development marked by the degradation of maternally provided mRNAs and initiation of zygotic gene expression. However the changes occurring at the protein level during this transition remain unclear. Here we conducted protein profiling throughout zebrafish embryogenesis using quantitative mass spectrometry integrating transcriptomics and translatomics datasets. Our data shows that unlike RNA changes protein changes are less dynamic. Further increases in protein levels correlate with mRNA translation whereas declines in protein levels do not suggesting active protein degradation processes. Interestingly proteins from pure zygotic genes are present at fertilization challenging existing mRNA based gene classifications. As a proof of concept we utilized CRISPR Cas13d to target znf281b mRNA a gene whose protein significantly accumulates within the first two hpf demonstrating its crucial role in development. Consequently our protein profiling coupled with CRISPR Cas13d offers a new approach to unravel maternal mRNAs function during embryonic development. Overall design: Two biological replicate samples containing 50 zebrafish oocytes per stage were collected by pairing females in natural matings to ''purge'' mature eggs and used to establish an oogenesis time line. Fish were euthanized and ovaries harvested within 1–11 days post purging dpp. Stage I and II oocytes were collected at 1–2 dpp stage III oocytes germinal vesicle in central position at 4–7 dpp and stage IV oocytes germinal vesicle asymmetrically located at 8–10 dpp. Oocyte isolations were based on73 and conducted in isolation medium Leibovitz L 15 Sigma Aldrich #L5520 plus Collagenase I and II depending on the stages as follows. Stages I and II were isolated with 3 mg/mL Collagenase I Sigma Aldrich C0130 and 3 mg/mL Collagenase II Gibco 17101015. Stage III was isolated with 3 mg/mL Collagenase I Sigma Aldrich C0130. Stage IV was isolated by mechanical stripping with forceps and nee…   pubmed:39302832   pCas13d al1 4hpf rep2 GSM8446428   source name:Whole embryo|tissue:Whole embryo|developmental stage:4hpf|genotype:AB TF and TLF|geo loc name:missing|collection date:missing pCas13d al1 4hpf rep2 Raw reads from zebrafish oocytes stages I IV and embryos 0 2 4 and 6 hpf were demultiplexed into FASTQ format allowing up to one mismatch using Illumina bcl convert 3.10.5. Reads were aligned to UCSC genome danRer11 with STAR aligner version 2.7.3a using Ensembl 106 gene models. Counts were converted to TPM values using RSEM version v1.3.0 and all subsequent analysis was done using TPM. Assembly: danRer11 Ensembl 106 gene models Supplementary files format and content: comma separated files with TPMs from all replicates Whole embryo   Isolated oocytes were snap frozen and RNA was extracted with TRIzol Invitrogen #15596026 following manufacturer’s protocols. 0 2 4 and 6 hpf zebrafish embryo RNA was extracted with Direct zol RNA MicroPrep kit Zymo Research #R2062 following manufacturer’s protocols. Zebrafish embryos 0 2 4 and 6 hpf total RNA sequencing libraries were generated from 500ng of high quality total RNA as assessed using Bioanalyzer Agilent. Libraries were made according to the manufacturer’s directions for the TruSeq Stranded Total RNA Library Prep Gold Illumina #20020598 and TruSeq RNA Single Indexes Sets A and B Illumina #20020492 and #20020493. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer Agilent Technologies and Qubit Fluorometer Life Technologies. Libraries were pooled quantified and sequenced as 75bp single reads on a high output flow cell using the Illumina NextSeq500 instrument. Following sequencing Illumina Primary Analysis version RTA 2.11.3.0 and bcl convert 3.10.5 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish oocytes stages I IV total RNA Seq libraries were generated from 100 ng of high quality total RNA as assessed by a Bioanalyzer Agilent Technologies. Libraries were prepared according to manufacturer’s instructions using the TruSeq Stranded Total RNA Library Prep Gold Illumina Cat. No. 20020598 and TruSeq RNA Single Indexes; Sets A and B Illumina Cat. No. 20020492 and 20020493. Resulting short fragment libraries were checked for quality and quantity using a Bioanalyzer and a Qubit Fluorometer Life Technologies. Libraries were normalized pooled and sequenced on a NextSeq 500 instrument Illumina as 75 bp single end reads on a high output flowcell using NextSeq Control Software 2.2.0.4. Following sequencing Illumina Primary Analysis version NextSeq RTA 2.4.11 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish znf281b knockdown embryos…   tissue:Whole embryo|developmental stage:4hpf|genotype:AB TF and TLF GSM8446428 GSM8446428: pCas13d al1 4hpf rep2; Danio rerio; RNA Seq GSM8446428 r1 GSM8446428 1 Isolated oocytes were snap frozen and RNA was extracted with TRIzol Invitrogen #15596026 following manufacturer's protocols. 0 2 4 and 6 hpf zebrafish embryo RNA was extracted with Direct zol RNA MicroPrep kit Zymo Research #R2062 following manufacturer's protocols. Zebrafish embryos 0 2 4 and 6 hpf total RNA sequencing libraries were generated from 500ng of high quality total RNA as assessed using Bioanalyzer Agilent. Libraries were made according to the manufacturer's directions for the TruSeq Stranded Total RNA Library Prep Gold Illumina #20020598 and TruSeq RNA Single Indexes Sets A and B Illumina #20020492 and #20020493. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer Agilent Technologies and Qubit Fluorometer Life Technologies. Libraries were pooled quantified and sequenced as 75bp single reads on a high output flow cell using the Illumina NextSeq500 instrument. Following sequencing Illumina Primary Analysis version RTA 2.11.3.0 and bcl convert 3.10.5 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish oocytes stages I IV total RNA Seq libraries were generated from 100 ng of high quality total RNA as assessed by a Bioanalyzer Agilent Technologies. Libraries were prepared according to manufacturer's instructions using the TruSeq Stranded Total RNA Library Prep Gold Illumina Cat. No. 20020598 and TruSeq RNA Single Indexes; Sets A and B Illumina Cat. No. 20020492 and 20020493. Resulting short fragment libraries were checked for quality and quantity using a Bioanalyzer and a Qubit Fluorometer Life Technologies. Libraries were normalized pooled and sequenced on a NextSeq 500 instrument Illumina as 75 bp single end reads on a high output flowcell using NextSeq Control Software 2.2.0.4. Following sequencing Illumina Primary Analysis version NextSeq RTA 2.4.11 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish znf281b knockdown embryos…   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP495323     L67854_S10_L001_R1_001.fastq.gz L67854_S10_L001_R2_001.fastq.gz fastq fastq 2528276500.0 25282765.0 GSM8446428 r1 0:50 1:50 A:664250318;C:589008342;G:598067975;T:676949865;N:0 50 50     664250318 589008342 598067975 676949865 0 SRX25634205 SRS22278605 SRA1942483 Computational Biology, Stowers Institute for Medical Research Computational Biology, Stowers Institute for Medical Research                       B B biological fallback assumption illumina nextseq unknown poly_a trueseq bulk bulk bulk   United States 2024-08-07 Blastula Embryo Whole Organism All anatomical structures
30751 30751 SRR30167502 SRX25634204 SRS22278602 SRP495323 PRJNA1088158 Protein profiling of zebrafish embryos unmasks regulatory layers during early embryogenesis. GSE261646 Transcriptome Analysis The maternal to zygotic transition is crucial in embryonic development marked by the degradation of maternally provided mRNAs and initiation of zygotic gene expression. However the changes occurring at the protein level during this transition remain unclear. Here we conducted protein profiling throughout zebrafish embryogenesis using quantitative mass spectrometry integrating transcriptomics and translatomics datasets. Our data shows that unlike RNA changes protein changes are less dynamic. Further increases in protein levels correlate with mRNA translation whereas declines in protein levels do not suggesting active protein degradation processes. Interestingly proteins from pure zygotic genes are present at fertilization challenging existing mRNA based gene classifications. As a proof of concept we utilized CRISPR Cas13d to target znf281b mRNA a gene whose protein significantly accumulates within the first two hpf demonstrating its crucial role in development. Consequently our protein profiling coupled with CRISPR Cas13d offers a new approach to unravel maternal mRNAs function during embryonic development. Overall design: Two biological replicate samples containing 50 zebrafish oocytes per stage were collected by pairing females in natural matings to ''purge'' mature eggs and used to establish an oogenesis time line. Fish were euthanized and ovaries harvested within 1–11 days post purging dpp. Stage I and II oocytes were collected at 1–2 dpp stage III oocytes germinal vesicle in central position at 4–7 dpp and stage IV oocytes germinal vesicle asymmetrically located at 8–10 dpp. Oocyte isolations were based on73 and conducted in isolation medium Leibovitz L 15 Sigma Aldrich #L5520 plus Collagenase I and II depending on the stages as follows. Stages I and II were isolated with 3 mg/mL Collagenase I Sigma Aldrich C0130 and 3 mg/mL Collagenase II Gibco 17101015. Stage III was isolated with 3 mg/mL Collagenase I Sigma Aldrich C0130. Stage IV was isolated by mechanical stripping with forceps and nee…   pubmed:39302832   pCas13d al1 4hpf rep1 GSM8446427   source name:Whole embryo|tissue:Whole embryo|developmental stage:4hpf|genotype:AB TF and TLF|geo loc name:missing|collection date:missing pCas13d al1 4hpf rep1 Raw reads from zebrafish oocytes stages I IV and embryos 0 2 4 and 6 hpf were demultiplexed into FASTQ format allowing up to one mismatch using Illumina bcl convert 3.10.5. Reads were aligned to UCSC genome danRer11 with STAR aligner version 2.7.3a using Ensembl 106 gene models. Counts were converted to TPM values using RSEM version v1.3.0 and all subsequent analysis was done using TPM. Assembly: danRer11 Ensembl 106 gene models Supplementary files format and content: comma separated files with TPMs from all replicates Whole embryo   Isolated oocytes were snap frozen and RNA was extracted with TRIzol Invitrogen #15596026 following manufacturer’s protocols. 0 2 4 and 6 hpf zebrafish embryo RNA was extracted with Direct zol RNA MicroPrep kit Zymo Research #R2062 following manufacturer’s protocols. Zebrafish embryos 0 2 4 and 6 hpf total RNA sequencing libraries were generated from 500ng of high quality total RNA as assessed using Bioanalyzer Agilent. Libraries were made according to the manufacturer’s directions for the TruSeq Stranded Total RNA Library Prep Gold Illumina #20020598 and TruSeq RNA Single Indexes Sets A and B Illumina #20020492 and #20020493. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer Agilent Technologies and Qubit Fluorometer Life Technologies. Libraries were pooled quantified and sequenced as 75bp single reads on a high output flow cell using the Illumina NextSeq500 instrument. Following sequencing Illumina Primary Analysis version RTA 2.11.3.0 and bcl convert 3.10.5 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish oocytes stages I IV total RNA Seq libraries were generated from 100 ng of high quality total RNA as assessed by a Bioanalyzer Agilent Technologies. Libraries were prepared according to manufacturer’s instructions using the TruSeq Stranded Total RNA Library Prep Gold Illumina Cat. No. 20020598 and TruSeq RNA Single Indexes; Sets A and B Illumina Cat. No. 20020492 and 20020493. Resulting short fragment libraries were checked for quality and quantity using a Bioanalyzer and a Qubit Fluorometer Life Technologies. Libraries were normalized pooled and sequenced on a NextSeq 500 instrument Illumina as 75 bp single end reads on a high output flowcell using NextSeq Control Software 2.2.0.4. Following sequencing Illumina Primary Analysis version NextSeq RTA 2.4.11 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish znf281b knockdown embryos…   tissue:Whole embryo|developmental stage:4hpf|genotype:AB TF and TLF GSM8446427 GSM8446427: pCas13d al1 4hpf rep1; Danio rerio; RNA Seq GSM8446427 r1 GSM8446427 1 Isolated oocytes were snap frozen and RNA was extracted with TRIzol Invitrogen #15596026 following manufacturer's protocols. 0 2 4 and 6 hpf zebrafish embryo RNA was extracted with Direct zol RNA MicroPrep kit Zymo Research #R2062 following manufacturer's protocols. Zebrafish embryos 0 2 4 and 6 hpf total RNA sequencing libraries were generated from 500ng of high quality total RNA as assessed using Bioanalyzer Agilent. Libraries were made according to the manufacturer's directions for the TruSeq Stranded Total RNA Library Prep Gold Illumina #20020598 and TruSeq RNA Single Indexes Sets A and B Illumina #20020492 and #20020493. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer Agilent Technologies and Qubit Fluorometer Life Technologies. Libraries were pooled quantified and sequenced as 75bp single reads on a high output flow cell using the Illumina NextSeq500 instrument. Following sequencing Illumina Primary Analysis version RTA 2.11.3.0 and bcl convert 3.10.5 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish oocytes stages I IV total RNA Seq libraries were generated from 100 ng of high quality total RNA as assessed by a Bioanalyzer Agilent Technologies. Libraries were prepared according to manufacturer's instructions using the TruSeq Stranded Total RNA Library Prep Gold Illumina Cat. No. 20020598 and TruSeq RNA Single Indexes; Sets A and B Illumina Cat. No. 20020492 and 20020493. Resulting short fragment libraries were checked for quality and quantity using a Bioanalyzer and a Qubit Fluorometer Life Technologies. Libraries were normalized pooled and sequenced on a NextSeq 500 instrument Illumina as 75 bp single end reads on a high output flowcell using NextSeq Control Software 2.2.0.4. Following sequencing Illumina Primary Analysis version NextSeq RTA 2.4.11 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish znf281b knockdown embryos…   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP495323     L67853_S9_L001_R1_001.fastq.gz L67853_S9_L001_R2_001.fastq.gz fastq fastq 2416897800.0 24168978.0 GSM8446427 r1 0:50 1:50 A:636467903;C:563622671;G:568978039;T:647829187;N:0 50 50     636467903 563622671 568978039 647829187 0 SRX25634204 SRS22278602 SRA1942483 Computational Biology, Stowers Institute for Medical Research Computational Biology, Stowers Institute for Medical Research                       B B biological fallback assumption illumina nextseq unknown poly_a trueseq bulk bulk bulk   United States 2024-08-07 Blastula Embryo Whole Organism All anatomical structures
30752 30752 SRR30167503 SRX25634203 SRS22278601 SRP495323 PRJNA1088158 Protein profiling of zebrafish embryos unmasks regulatory layers during early embryogenesis. GSE261646 Transcriptome Analysis The maternal to zygotic transition is crucial in embryonic development marked by the degradation of maternally provided mRNAs and initiation of zygotic gene expression. However the changes occurring at the protein level during this transition remain unclear. Here we conducted protein profiling throughout zebrafish embryogenesis using quantitative mass spectrometry integrating transcriptomics and translatomics datasets. Our data shows that unlike RNA changes protein changes are less dynamic. Further increases in protein levels correlate with mRNA translation whereas declines in protein levels do not suggesting active protein degradation processes. Interestingly proteins from pure zygotic genes are present at fertilization challenging existing mRNA based gene classifications. As a proof of concept we utilized CRISPR Cas13d to target znf281b mRNA a gene whose protein significantly accumulates within the first two hpf demonstrating its crucial role in development. Consequently our protein profiling coupled with CRISPR Cas13d offers a new approach to unravel maternal mRNAs function during embryonic development. Overall design: Two biological replicate samples containing 50 zebrafish oocytes per stage were collected by pairing females in natural matings to ''purge'' mature eggs and used to establish an oogenesis time line. Fish were euthanized and ovaries harvested within 1–11 days post purging dpp. Stage I and II oocytes were collected at 1–2 dpp stage III oocytes germinal vesicle in central position at 4–7 dpp and stage IV oocytes germinal vesicle asymmetrically located at 8–10 dpp. Oocyte isolations were based on73 and conducted in isolation medium Leibovitz L 15 Sigma Aldrich #L5520 plus Collagenase I and II depending on the stages as follows. Stages I and II were isolated with 3 mg/mL Collagenase I Sigma Aldrich C0130 and 3 mg/mL Collagenase II Gibco 17101015. Stage III was isolated with 3 mg/mL Collagenase I Sigma Aldrich C0130. Stage IV was isolated by mechanical stripping with forceps and nee…   pubmed:39302832   znf281b KD 4hpf rep3 GSM8446426   source name:Whole embryo|tissue:Whole embryo|developmental stage:4hpf|genotype:AB TF and TLF|geo loc name:missing|collection date:missing znf281b KD 4hpf rep3 Raw reads from zebrafish oocytes stages I IV and embryos 0 2 4 and 6 hpf were demultiplexed into FASTQ format allowing up to one mismatch using Illumina bcl convert 3.10.5. Reads were aligned to UCSC genome danRer11 with STAR aligner version 2.7.3a using Ensembl 106 gene models. Counts were converted to TPM values using RSEM version v1.3.0 and all subsequent analysis was done using TPM. Assembly: danRer11 Ensembl 106 gene models Supplementary files format and content: comma separated files with TPMs from all replicates Whole embryo   Isolated oocytes were snap frozen and RNA was extracted with TRIzol Invitrogen #15596026 following manufacturer’s protocols. 0 2 4 and 6 hpf zebrafish embryo RNA was extracted with Direct zol RNA MicroPrep kit Zymo Research #R2062 following manufacturer’s protocols. Zebrafish embryos 0 2 4 and 6 hpf total RNA sequencing libraries were generated from 500ng of high quality total RNA as assessed using Bioanalyzer Agilent. Libraries were made according to the manufacturer’s directions for the TruSeq Stranded Total RNA Library Prep Gold Illumina #20020598 and TruSeq RNA Single Indexes Sets A and B Illumina #20020492 and #20020493. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer Agilent Technologies and Qubit Fluorometer Life Technologies. Libraries were pooled quantified and sequenced as 75bp single reads on a high output flow cell using the Illumina NextSeq500 instrument. Following sequencing Illumina Primary Analysis version RTA 2.11.3.0 and bcl convert 3.10.5 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish oocytes stages I IV total RNA Seq libraries were generated from 100 ng of high quality total RNA as assessed by a Bioanalyzer Agilent Technologies. Libraries were prepared according to manufacturer’s instructions using the TruSeq Stranded Total RNA Library Prep Gold Illumina Cat. No. 20020598 and TruSeq RNA Single Indexes; Sets A and B Illumina Cat. No. 20020492 and 20020493. Resulting short fragment libraries were checked for quality and quantity using a Bioanalyzer and a Qubit Fluorometer Life Technologies. Libraries were normalized pooled and sequenced on a NextSeq 500 instrument Illumina as 75 bp single end reads on a high output flowcell using NextSeq Control Software 2.2.0.4. Following sequencing Illumina Primary Analysis version NextSeq RTA 2.4.11 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish znf281b knockdown embryos…   tissue:Whole embryo|developmental stage:4hpf|genotype:AB TF and TLF GSM8446426 GSM8446426: znf281b KD 4hpf rep3; Danio rerio; RNA Seq GSM8446426 r1 GSM8446426 1 Isolated oocytes were snap frozen and RNA was extracted with TRIzol Invitrogen #15596026 following manufacturer's protocols. 0 2 4 and 6 hpf zebrafish embryo RNA was extracted with Direct zol RNA MicroPrep kit Zymo Research #R2062 following manufacturer's protocols. Zebrafish embryos 0 2 4 and 6 hpf total RNA sequencing libraries were generated from 500ng of high quality total RNA as assessed using Bioanalyzer Agilent. Libraries were made according to the manufacturer's directions for the TruSeq Stranded Total RNA Library Prep Gold Illumina #20020598 and TruSeq RNA Single Indexes Sets A and B Illumina #20020492 and #20020493. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer Agilent Technologies and Qubit Fluorometer Life Technologies. Libraries were pooled quantified and sequenced as 75bp single reads on a high output flow cell using the Illumina NextSeq500 instrument. Following sequencing Illumina Primary Analysis version RTA 2.11.3.0 and bcl convert 3.10.5 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish oocytes stages I IV total RNA Seq libraries were generated from 100 ng of high quality total RNA as assessed by a Bioanalyzer Agilent Technologies. Libraries were prepared according to manufacturer's instructions using the TruSeq Stranded Total RNA Library Prep Gold Illumina Cat. No. 20020598 and TruSeq RNA Single Indexes; Sets A and B Illumina Cat. No. 20020492 and 20020493. Resulting short fragment libraries were checked for quality and quantity using a Bioanalyzer and a Qubit Fluorometer Life Technologies. Libraries were normalized pooled and sequenced on a NextSeq 500 instrument Illumina as 75 bp single end reads on a high output flowcell using NextSeq Control Software 2.2.0.4. Following sequencing Illumina Primary Analysis version NextSeq RTA 2.4.11 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish znf281b knockdown embryos…   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP495323     L67851_S7_L001_R1_001.fastq.gz L67851_S7_L001_R2_001.fastq.gz fastq fastq 2366597500.0 23665975.0 GSM8446426 r1 0:50 1:50 A:620883932;C:547989723;G:563335901;T:634387944;N:0 50 50     620883932 547989723 563335901 634387944 0 SRX25634203 SRS22278601 SRA1942483 Computational Biology, Stowers Institute for Medical Research Computational Biology, Stowers Institute for Medical Research                       B B biological fallback assumption illumina nextseq unknown poly_a trueseq bulk bulk bulk   United States 2024-08-07 Blastula Embryo Whole Organism All anatomical structures
30753 30753 SRR30167504 SRX25634202 SRS22278600 SRP495323 PRJNA1088158 Protein profiling of zebrafish embryos unmasks regulatory layers during early embryogenesis. GSE261646 Transcriptome Analysis The maternal to zygotic transition is crucial in embryonic development marked by the degradation of maternally provided mRNAs and initiation of zygotic gene expression. However the changes occurring at the protein level during this transition remain unclear. Here we conducted protein profiling throughout zebrafish embryogenesis using quantitative mass spectrometry integrating transcriptomics and translatomics datasets. Our data shows that unlike RNA changes protein changes are less dynamic. Further increases in protein levels correlate with mRNA translation whereas declines in protein levels do not suggesting active protein degradation processes. Interestingly proteins from pure zygotic genes are present at fertilization challenging existing mRNA based gene classifications. As a proof of concept we utilized CRISPR Cas13d to target znf281b mRNA a gene whose protein significantly accumulates within the first two hpf demonstrating its crucial role in development. Consequently our protein profiling coupled with CRISPR Cas13d offers a new approach to unravel maternal mRNAs function during embryonic development. Overall design: Two biological replicate samples containing 50 zebrafish oocytes per stage were collected by pairing females in natural matings to ''purge'' mature eggs and used to establish an oogenesis time line. Fish were euthanized and ovaries harvested within 1–11 days post purging dpp. Stage I and II oocytes were collected at 1–2 dpp stage III oocytes germinal vesicle in central position at 4–7 dpp and stage IV oocytes germinal vesicle asymmetrically located at 8–10 dpp. Oocyte isolations were based on73 and conducted in isolation medium Leibovitz L 15 Sigma Aldrich #L5520 plus Collagenase I and II depending on the stages as follows. Stages I and II were isolated with 3 mg/mL Collagenase I Sigma Aldrich C0130 and 3 mg/mL Collagenase II Gibco 17101015. Stage III was isolated with 3 mg/mL Collagenase I Sigma Aldrich C0130. Stage IV was isolated by mechanical stripping with forceps and nee…   pubmed:39302832   znf281b KD 4hpf rep2 GSM8446425   source name:Whole embryo|tissue:Whole embryo|developmental stage:4hpf|genotype:AB TF and TLF|geo loc name:missing|collection date:missing znf281b KD 4hpf rep2 Raw reads from zebrafish oocytes stages I IV and embryos 0 2 4 and 6 hpf were demultiplexed into FASTQ format allowing up to one mismatch using Illumina bcl convert 3.10.5. Reads were aligned to UCSC genome danRer11 with STAR aligner version 2.7.3a using Ensembl 106 gene models. Counts were converted to TPM values using RSEM version v1.3.0 and all subsequent analysis was done using TPM. Assembly: danRer11 Ensembl 106 gene models Supplementary files format and content: comma separated files with TPMs from all replicates Whole embryo   Isolated oocytes were snap frozen and RNA was extracted with TRIzol Invitrogen #15596026 following manufacturer’s protocols. 0 2 4 and 6 hpf zebrafish embryo RNA was extracted with Direct zol RNA MicroPrep kit Zymo Research #R2062 following manufacturer’s protocols. Zebrafish embryos 0 2 4 and 6 hpf total RNA sequencing libraries were generated from 500ng of high quality total RNA as assessed using Bioanalyzer Agilent. Libraries were made according to the manufacturer’s directions for the TruSeq Stranded Total RNA Library Prep Gold Illumina #20020598 and TruSeq RNA Single Indexes Sets A and B Illumina #20020492 and #20020493. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer Agilent Technologies and Qubit Fluorometer Life Technologies. Libraries were pooled quantified and sequenced as 75bp single reads on a high output flow cell using the Illumina NextSeq500 instrument. Following sequencing Illumina Primary Analysis version RTA 2.11.3.0 and bcl convert 3.10.5 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish oocytes stages I IV total RNA Seq libraries were generated from 100 ng of high quality total RNA as assessed by a Bioanalyzer Agilent Technologies. Libraries were prepared according to manufacturer’s instructions using the TruSeq Stranded Total RNA Library Prep Gold Illumina Cat. No. 20020598 and TruSeq RNA Single Indexes; Sets A and B Illumina Cat. No. 20020492 and 20020493. Resulting short fragment libraries were checked for quality and quantity using a Bioanalyzer and a Qubit Fluorometer Life Technologies. Libraries were normalized pooled and sequenced on a NextSeq 500 instrument Illumina as 75 bp single end reads on a high output flowcell using NextSeq Control Software 2.2.0.4. Following sequencing Illumina Primary Analysis version NextSeq RTA 2.4.11 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish znf281b knockdown embryos…   tissue:Whole embryo|developmental stage:4hpf|genotype:AB TF and TLF GSM8446425 GSM8446425: znf281b KD 4hpf rep2; Danio rerio; RNA Seq GSM8446425 r1 GSM8446425 1 Isolated oocytes were snap frozen and RNA was extracted with TRIzol Invitrogen #15596026 following manufacturer's protocols. 0 2 4 and 6 hpf zebrafish embryo RNA was extracted with Direct zol RNA MicroPrep kit Zymo Research #R2062 following manufacturer's protocols. Zebrafish embryos 0 2 4 and 6 hpf total RNA sequencing libraries were generated from 500ng of high quality total RNA as assessed using Bioanalyzer Agilent. Libraries were made according to the manufacturer's directions for the TruSeq Stranded Total RNA Library Prep Gold Illumina #20020598 and TruSeq RNA Single Indexes Sets A and B Illumina #20020492 and #20020493. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer Agilent Technologies and Qubit Fluorometer Life Technologies. Libraries were pooled quantified and sequenced as 75bp single reads on a high output flow cell using the Illumina NextSeq500 instrument. Following sequencing Illumina Primary Analysis version RTA 2.11.3.0 and bcl convert 3.10.5 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish oocytes stages I IV total RNA Seq libraries were generated from 100 ng of high quality total RNA as assessed by a Bioanalyzer Agilent Technologies. Libraries were prepared according to manufacturer's instructions using the TruSeq Stranded Total RNA Library Prep Gold Illumina Cat. No. 20020598 and TruSeq RNA Single Indexes; Sets A and B Illumina Cat. No. 20020492 and 20020493. Resulting short fragment libraries were checked for quality and quantity using a Bioanalyzer and a Qubit Fluorometer Life Technologies. Libraries were normalized pooled and sequenced on a NextSeq 500 instrument Illumina as 75 bp single end reads on a high output flowcell using NextSeq Control Software 2.2.0.4. Following sequencing Illumina Primary Analysis version NextSeq RTA 2.4.11 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish znf281b knockdown embryos…   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP495323     L67850_S6_L001_R1_001.fastq.gz L67850_S6_L001_R2_001.fastq.gz fastq fastq 2497293900.0 24972939.0 GSM8446425 r1 0:50 1:50 A:657452880;C:579618079;G:589789328;T:670433613;N:0 50 50     657452880 579618079 589789328 670433613 0 SRX25634202 SRS22278600 SRA1942483 Computational Biology, Stowers Institute for Medical Research Computational Biology, Stowers Institute for Medical Research                       B B biological fallback assumption illumina nextseq unknown poly_a trueseq bulk bulk bulk   United States 2024-08-07 Blastula Embryo Whole Organism All anatomical structures
30754 30754 SRR30167505 SRX25634201 SRS22278596 SRP495323 PRJNA1088158 Protein profiling of zebrafish embryos unmasks regulatory layers during early embryogenesis. GSE261646 Transcriptome Analysis The maternal to zygotic transition is crucial in embryonic development marked by the degradation of maternally provided mRNAs and initiation of zygotic gene expression. However the changes occurring at the protein level during this transition remain unclear. Here we conducted protein profiling throughout zebrafish embryogenesis using quantitative mass spectrometry integrating transcriptomics and translatomics datasets. Our data shows that unlike RNA changes protein changes are less dynamic. Further increases in protein levels correlate with mRNA translation whereas declines in protein levels do not suggesting active protein degradation processes. Interestingly proteins from pure zygotic genes are present at fertilization challenging existing mRNA based gene classifications. As a proof of concept we utilized CRISPR Cas13d to target znf281b mRNA a gene whose protein significantly accumulates within the first two hpf demonstrating its crucial role in development. Consequently our protein profiling coupled with CRISPR Cas13d offers a new approach to unravel maternal mRNAs function during embryonic development. Overall design: Two biological replicate samples containing 50 zebrafish oocytes per stage were collected by pairing females in natural matings to ''purge'' mature eggs and used to establish an oogenesis time line. Fish were euthanized and ovaries harvested within 1–11 days post purging dpp. Stage I and II oocytes were collected at 1–2 dpp stage III oocytes germinal vesicle in central position at 4–7 dpp and stage IV oocytes germinal vesicle asymmetrically located at 8–10 dpp. Oocyte isolations were based on73 and conducted in isolation medium Leibovitz L 15 Sigma Aldrich #L5520 plus Collagenase I and II depending on the stages as follows. Stages I and II were isolated with 3 mg/mL Collagenase I Sigma Aldrich C0130 and 3 mg/mL Collagenase II Gibco 17101015. Stage III was isolated with 3 mg/mL Collagenase I Sigma Aldrich C0130. Stage IV was isolated by mechanical stripping with forceps and nee…   pubmed:39302832   znf281b KD 4hpf rep1 GSM8446424   source name:Whole embryo|tissue:Whole embryo|developmental stage:4hpf|genotype:AB TF and TLF|geo loc name:missing|collection date:missing znf281b KD 4hpf rep1 Raw reads from zebrafish oocytes stages I IV and embryos 0 2 4 and 6 hpf were demultiplexed into FASTQ format allowing up to one mismatch using Illumina bcl convert 3.10.5. Reads were aligned to UCSC genome danRer11 with STAR aligner version 2.7.3a using Ensembl 106 gene models. Counts were converted to TPM values using RSEM version v1.3.0 and all subsequent analysis was done using TPM. Assembly: danRer11 Ensembl 106 gene models Supplementary files format and content: comma separated files with TPMs from all replicates Whole embryo   Isolated oocytes were snap frozen and RNA was extracted with TRIzol Invitrogen #15596026 following manufacturer’s protocols. 0 2 4 and 6 hpf zebrafish embryo RNA was extracted with Direct zol RNA MicroPrep kit Zymo Research #R2062 following manufacturer’s protocols. Zebrafish embryos 0 2 4 and 6 hpf total RNA sequencing libraries were generated from 500ng of high quality total RNA as assessed using Bioanalyzer Agilent. Libraries were made according to the manufacturer’s directions for the TruSeq Stranded Total RNA Library Prep Gold Illumina #20020598 and TruSeq RNA Single Indexes Sets A and B Illumina #20020492 and #20020493. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer Agilent Technologies and Qubit Fluorometer Life Technologies. Libraries were pooled quantified and sequenced as 75bp single reads on a high output flow cell using the Illumina NextSeq500 instrument. Following sequencing Illumina Primary Analysis version RTA 2.11.3.0 and bcl convert 3.10.5 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish oocytes stages I IV total RNA Seq libraries were generated from 100 ng of high quality total RNA as assessed by a Bioanalyzer Agilent Technologies. Libraries were prepared according to manufacturer’s instructions using the TruSeq Stranded Total RNA Library Prep Gold Illumina Cat. No. 20020598 and TruSeq RNA Single Indexes; Sets A and B Illumina Cat. No. 20020492 and 20020493. Resulting short fragment libraries were checked for quality and quantity using a Bioanalyzer and a Qubit Fluorometer Life Technologies. Libraries were normalized pooled and sequenced on a NextSeq 500 instrument Illumina as 75 bp single end reads on a high output flowcell using NextSeq Control Software 2.2.0.4. Following sequencing Illumina Primary Analysis version NextSeq RTA 2.4.11 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish znf281b knockdown embryos…   tissue:Whole embryo|developmental stage:4hpf|genotype:AB TF and TLF GSM8446424 GSM8446424: znf281b KD 4hpf rep1; Danio rerio; RNA Seq GSM8446424 r1 GSM8446424 1 Isolated oocytes were snap frozen and RNA was extracted with TRIzol Invitrogen #15596026 following manufacturer's protocols. 0 2 4 and 6 hpf zebrafish embryo RNA was extracted with Direct zol RNA MicroPrep kit Zymo Research #R2062 following manufacturer's protocols. Zebrafish embryos 0 2 4 and 6 hpf total RNA sequencing libraries were generated from 500ng of high quality total RNA as assessed using Bioanalyzer Agilent. Libraries were made according to the manufacturer's directions for the TruSeq Stranded Total RNA Library Prep Gold Illumina #20020598 and TruSeq RNA Single Indexes Sets A and B Illumina #20020492 and #20020493. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer Agilent Technologies and Qubit Fluorometer Life Technologies. Libraries were pooled quantified and sequenced as 75bp single reads on a high output flow cell using the Illumina NextSeq500 instrument. Following sequencing Illumina Primary Analysis version RTA 2.11.3.0 and bcl convert 3.10.5 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish oocytes stages I IV total RNA Seq libraries were generated from 100 ng of high quality total RNA as assessed by a Bioanalyzer Agilent Technologies. Libraries were prepared according to manufacturer's instructions using the TruSeq Stranded Total RNA Library Prep Gold Illumina Cat. No. 20020598 and TruSeq RNA Single Indexes; Sets A and B Illumina Cat. No. 20020492 and 20020493. Resulting short fragment libraries were checked for quality and quantity using a Bioanalyzer and a Qubit Fluorometer Life Technologies. Libraries were normalized pooled and sequenced on a NextSeq 500 instrument Illumina as 75 bp single end reads on a high output flowcell using NextSeq Control Software 2.2.0.4. Following sequencing Illumina Primary Analysis version NextSeq RTA 2.4.11 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish znf281b knockdown embryos…   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP495323     L67849_S5_L001_R1_001.fastq.gz L67849_S5_L001_R2_001.fastq.gz fastq fastq 2607154500.0 26071545.0 GSM8446424 r1 0:50 1:50 A:685092511;C:605305852;G:618493032;T:698263105;N:0 50 50     685092511 605305852 618493032 698263105 0 SRX25634201 SRS22278596 SRA1942483 Computational Biology, Stowers Institute for Medical Research Computational Biology, Stowers Institute for Medical Research                       B B biological fallback assumption illumina nextseq unknown poly_a trueseq bulk bulk bulk   United States 2024-08-07 Blastula Embryo Whole Organism All anatomical structures
30766 30766 SRR28348932 SRX23954964 SRS20755385 SRP495323 PRJNA1088158 Protein profiling of zebrafish embryos unmasks regulatory layers during early embryogenesis. GSE261646 Transcriptome Analysis The maternal to zygotic transition is crucial in embryonic development marked by the degradation of maternally provided mRNAs and initiation of zygotic gene expression. However the changes occurring at the protein level during this transition remain unclear. Here we conducted protein profiling throughout zebrafish embryogenesis using quantitative mass spectrometry integrating transcriptomics and translatomics datasets. Our data shows that unlike RNA changes protein changes are less dynamic. Further increases in protein levels correlate with mRNA translation whereas declines in protein levels do not suggesting active protein degradation processes. Interestingly proteins from pure zygotic genes are present at fertilization challenging existing mRNA based gene classifications. As a proof of concept we utilized CRISPR Cas13d to target znf281b mRNA a gene whose protein significantly accumulates within the first two hpf demonstrating its crucial role in development. Consequently our protein profiling coupled with CRISPR Cas13d offers a new approach to unravel maternal mRNAs function during embryonic development. Overall design: Two biological replicate samples containing 50 zebrafish oocytes per stage were collected by pairing females in natural matings to ''purge'' mature eggs and used to establish an oogenesis time line. Fish were euthanized and ovaries harvested within 1–11 days post purging dpp. Stage I and II oocytes were collected at 1–2 dpp stage III oocytes germinal vesicle in central position at 4–7 dpp and stage IV oocytes germinal vesicle asymmetrically located at 8–10 dpp. Oocyte isolations were based on73 and conducted in isolation medium Leibovitz L 15 Sigma Aldrich #L5520 plus Collagenase I and II depending on the stages as follows. Stages I and II were isolated with 3 mg/mL Collagenase I Sigma Aldrich C0130 and 3 mg/mL Collagenase II Gibco 17101015. Stage III was isolated with 3 mg/mL Collagenase I Sigma Aldrich C0130. Stage IV was isolated by mechanical stripping with forceps and nee…   pubmed:39302832   Zebrafish Embryo 4hpf rep3 GSM8147861   source name:Whole embryo|tissue:Whole embryo|developmental stage:4hpf|genotype:AB TF and TLF|geo loc name:missing|collection date:missing Zebrafish Embryo 4hpf rep3 Raw reads from zebrafish oocytes stages I IV and embryos 0 2 4 and 6 hpf were demultiplexed into FASTQ format allowing up to one mismatch using Illumina bcl convert 3.10.5. Reads were aligned to UCSC genome danRer11 with STAR aligner version 2.7.3a using Ensembl 106 gene models. Counts were converted to TPM values using RSEM version v1.3.0 and all subsequent analysis was done using TPM. Assembly: danRer11 Ensembl 106 gene models Supplementary files format and content: comma separated files with TPMs from all replicates Whole embryo   Isolated oocytes were snap frozen and RNA was extracted with TRIzol Invitrogen #15596026 following manufacturer’s protocols. 0 2 4 and 6 hpf zebrafish embryo RNA was extracted with Direct zol RNA MicroPrep kit Zymo Research #R2062 following manufacturer’s protocols. Ribo dep Stranded RNA Seq Zebrafish embryos 0 2 4 and 6 hpf total RNA sequencing libraries were generated from 500ng of high quality total RNA as assessed using Bioanalyzer Agilent. Libraries were made according to the manufacturer’s directions for the TruSeq Stranded Total RNA Library Prep Gold Illumina #20020598 and TruSeq RNA Single Indexes Sets A and B Illumina #20020492 and #20020493. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer Agilent Technologies and Qubit Fluorometer Life Technologies. Libraries were pooled quantified and sequenced as 75bp single reads on a high output flow cell using the Illumina NextSeq500 instrument. Following sequencing Illumina Primary Analysis version RTA 2.11.3.0 and bcl convert 3.10.5 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish oocytes stages I IV total RNA Seq libraries were generated from 100 ng of high quality total RNA as assessed by a Bioanalyzer Agilent Technologies. Libraries were prepared according to manufacturer’s instructions using the TruSeq Stranded Total RNA Library Prep Gold Illumina Cat. No. 20020598 and TruSeq RNA Single Indexes; Sets A and B Illumina Cat. No. 20020492 and 20020493. Resulting short fragment libraries were checked for quality and quantity using a Bioanalyzer and a Qubit Fluorometer Life Technologies. Libraries were normalized pooled and sequenced on a NextSeq 500 instrument Illumina as 75 bp single end reads on a high output flowcell using NextSeq Control Software 2.2.0.4. Following sequencing Illumina Primary Analysis version NextSeq RTA 2.4.11 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files.   tissue:Whole embryo|developmental stage:4hpf|genotype:AB TF and TLF GSM8147861 GSM8147861: Zebrafish Embryo 4hpf rep3; Danio rerio; RNA Seq GSM8147861 r1 GSM8147861 1 Isolated oocytes were snap frozen and RNA was extracted with TRIzol Invitrogen #15596026 following manufacturer's protocols. 0 2 4 and 6 hpf zebrafish embryo RNA was extracted with Direct zol RNA MicroPrep kit Zymo Research #R2062 following manufacturer's protocols. Ribo dep Stranded RNA Seq Zebrafish embryos 0 2 4 and 6 hpf total RNA sequencing libraries were generated from 500ng of high quality total RNA as assessed using Bioanalyzer Agilent. Libraries were made according to the manufacturer's directions for the TruSeq Stranded Total RNA Library Prep Gold Illumina #20020598 and TruSeq RNA Single Indexes Sets A and B Illumina #20020492 and #20020493. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer Agilent Technologies and Qubit Fluorometer Life Technologies. Libraries were pooled quantified and sequenced as 75bp single reads on a high output flow cell using the Illumina NextSeq500 instrument. Following sequencing Illumina Primary Analysis version RTA 2.11.3.0 and bcl convert 3.10.5 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish oocytes stages I IV total RNA Seq libraries were generated from 100 ng of high quality total RNA as assessed by a Bioanalyzer Agilent Technologies. Libraries were prepared according to manufacturer's instructions using the TruSeq Stranded Total RNA Library Prep Gold Illumina Cat. No. 20020598 and TruSeq RNA Single Indexes; Sets A and B Illumina Cat. No. 20020492 and 20020493. Resulting short fragment libraries were checked for quality and quantity using a Bioanalyzer and a Qubit Fluorometer Life Technologies. Libraries were normalized pooled and sequenced on a NextSeq 500 instrument Illumina as 75 bp single end reads on a high output flowcell using NextSeq Control Software 2.2.0.4. Following sequencing Illumina Primary Analysis version NextSeq RTA 2.4.11 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files.   RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA NextSeq 500   SRP495323     s_4hpf_3.fastq fastq 1795835160.0 23629410.0 GSM8147861 r1 0:76 A:438577940;C:442051089;G:414188325;T:500933940;N:83866 76       438577940 442051089 414188325 500933940 83866 SRX23954964 SRS20755385 SRA1824599 Computational Biology, Stowers Institute for Medical Research Computational Biology, Stowers Institute for Medical Research                       B   usable mapping rate illumina nextseq unknown cdna_unspecified trueseq bulk bulk bulk   United States 2024-03-14 Blastula Embryo Whole Organism All anatomical structures
30767 30767 SRR28348933 SRX23954963 SRS20755384 SRP495323 PRJNA1088158 Protein profiling of zebrafish embryos unmasks regulatory layers during early embryogenesis. GSE261646 Transcriptome Analysis The maternal to zygotic transition is crucial in embryonic development marked by the degradation of maternally provided mRNAs and initiation of zygotic gene expression. However the changes occurring at the protein level during this transition remain unclear. Here we conducted protein profiling throughout zebrafish embryogenesis using quantitative mass spectrometry integrating transcriptomics and translatomics datasets. Our data shows that unlike RNA changes protein changes are less dynamic. Further increases in protein levels correlate with mRNA translation whereas declines in protein levels do not suggesting active protein degradation processes. Interestingly proteins from pure zygotic genes are present at fertilization challenging existing mRNA based gene classifications. As a proof of concept we utilized CRISPR Cas13d to target znf281b mRNA a gene whose protein significantly accumulates within the first two hpf demonstrating its crucial role in development. Consequently our protein profiling coupled with CRISPR Cas13d offers a new approach to unravel maternal mRNAs function during embryonic development. Overall design: Two biological replicate samples containing 50 zebrafish oocytes per stage were collected by pairing females in natural matings to ''purge'' mature eggs and used to establish an oogenesis time line. Fish were euthanized and ovaries harvested within 1–11 days post purging dpp. Stage I and II oocytes were collected at 1–2 dpp stage III oocytes germinal vesicle in central position at 4–7 dpp and stage IV oocytes germinal vesicle asymmetrically located at 8–10 dpp. Oocyte isolations were based on73 and conducted in isolation medium Leibovitz L 15 Sigma Aldrich #L5520 plus Collagenase I and II depending on the stages as follows. Stages I and II were isolated with 3 mg/mL Collagenase I Sigma Aldrich C0130 and 3 mg/mL Collagenase II Gibco 17101015. Stage III was isolated with 3 mg/mL Collagenase I Sigma Aldrich C0130. Stage IV was isolated by mechanical stripping with forceps and nee…   pubmed:39302832   Zebrafish Embryo 4hpf rep2 GSM8147860   source name:Whole embryo|tissue:Whole embryo|developmental stage:4hpf|genotype:AB TF and TLF|geo loc name:missing|collection date:missing Zebrafish Embryo 4hpf rep2 Raw reads from zebrafish oocytes stages I IV and embryos 0 2 4 and 6 hpf were demultiplexed into FASTQ format allowing up to one mismatch using Illumina bcl convert 3.10.5. Reads were aligned to UCSC genome danRer11 with STAR aligner version 2.7.3a using Ensembl 106 gene models. Counts were converted to TPM values using RSEM version v1.3.0 and all subsequent analysis was done using TPM. Assembly: danRer11 Ensembl 106 gene models Supplementary files format and content: comma separated files with TPMs from all replicates Whole embryo   Isolated oocytes were snap frozen and RNA was extracted with TRIzol Invitrogen #15596026 following manufacturer’s protocols. 0 2 4 and 6 hpf zebrafish embryo RNA was extracted with Direct zol RNA MicroPrep kit Zymo Research #R2062 following manufacturer’s protocols. Ribo dep Stranded RNA Seq Zebrafish embryos 0 2 4 and 6 hpf total RNA sequencing libraries were generated from 500ng of high quality total RNA as assessed using Bioanalyzer Agilent. Libraries were made according to the manufacturer’s directions for the TruSeq Stranded Total RNA Library Prep Gold Illumina #20020598 and TruSeq RNA Single Indexes Sets A and B Illumina #20020492 and #20020493. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer Agilent Technologies and Qubit Fluorometer Life Technologies. Libraries were pooled quantified and sequenced as 75bp single reads on a high output flow cell using the Illumina NextSeq500 instrument. Following sequencing Illumina Primary Analysis version RTA 2.11.3.0 and bcl convert 3.10.5 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish oocytes stages I IV total RNA Seq libraries were generated from 100 ng of high quality total RNA as assessed by a Bioanalyzer Agilent Technologies. Libraries were prepared according to manufacturer’s instructions using the TruSeq Stranded Total RNA Library Prep Gold Illumina Cat. No. 20020598 and TruSeq RNA Single Indexes; Sets A and B Illumina Cat. No. 20020492 and 20020493. Resulting short fragment libraries were checked for quality and quantity using a Bioanalyzer and a Qubit Fluorometer Life Technologies. Libraries were normalized pooled and sequenced on a NextSeq 500 instrument Illumina as 75 bp single end reads on a high output flowcell using NextSeq Control Software 2.2.0.4. Following sequencing Illumina Primary Analysis version NextSeq RTA 2.4.11 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files.   tissue:Whole embryo|developmental stage:4hpf|genotype:AB TF and TLF GSM8147860 GSM8147860: Zebrafish Embryo 4hpf rep2; Danio rerio; RNA Seq GSM8147860 r1 GSM8147860 1 Isolated oocytes were snap frozen and RNA was extracted with TRIzol Invitrogen #15596026 following manufacturer's protocols. 0 2 4 and 6 hpf zebrafish embryo RNA was extracted with Direct zol RNA MicroPrep kit Zymo Research #R2062 following manufacturer's protocols. Ribo dep Stranded RNA Seq Zebrafish embryos 0 2 4 and 6 hpf total RNA sequencing libraries were generated from 500ng of high quality total RNA as assessed using Bioanalyzer Agilent. Libraries were made according to the manufacturer's directions for the TruSeq Stranded Total RNA Library Prep Gold Illumina #20020598 and TruSeq RNA Single Indexes Sets A and B Illumina #20020492 and #20020493. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer Agilent Technologies and Qubit Fluorometer Life Technologies. Libraries were pooled quantified and sequenced as 75bp single reads on a high output flow cell using the Illumina NextSeq500 instrument. Following sequencing Illumina Primary Analysis version RTA 2.11.3.0 and bcl convert 3.10.5 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish oocytes stages I IV total RNA Seq libraries were generated from 100 ng of high quality total RNA as assessed by a Bioanalyzer Agilent Technologies. Libraries were prepared according to manufacturer's instructions using the TruSeq Stranded Total RNA Library Prep Gold Illumina Cat. No. 20020598 and TruSeq RNA Single Indexes; Sets A and B Illumina Cat. No. 20020492 and 20020493. Resulting short fragment libraries were checked for quality and quantity using a Bioanalyzer and a Qubit Fluorometer Life Technologies. Libraries were normalized pooled and sequenced on a NextSeq 500 instrument Illumina as 75 bp single end reads on a high output flowcell using NextSeq Control Software 2.2.0.4. Following sequencing Illumina Primary Analysis version NextSeq RTA 2.4.11 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files.   RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA NextSeq 500   SRP495323     s_4hpf_2.fastq fastq 1665073892.0 21908867.0 GSM8147860 r1 0:76 A:405272447;C:411954373;G:384597098;T:463177801;N:72173 76       405272447 411954373 384597098 463177801 72173 SRX23954963 SRS20755384 SRA1824599 Computational Biology, Stowers Institute for Medical Research Computational Biology, Stowers Institute for Medical Research                       B   usable mapping rate illumina nextseq unknown cdna_unspecified trueseq bulk bulk bulk   United States 2024-03-14 Blastula Embryo Whole Organism All anatomical structures
30768 30768 SRR28348934 SRX23954962 SRS20755383 SRP495323 PRJNA1088158 Protein profiling of zebrafish embryos unmasks regulatory layers during early embryogenesis. GSE261646 Transcriptome Analysis The maternal to zygotic transition is crucial in embryonic development marked by the degradation of maternally provided mRNAs and initiation of zygotic gene expression. However the changes occurring at the protein level during this transition remain unclear. Here we conducted protein profiling throughout zebrafish embryogenesis using quantitative mass spectrometry integrating transcriptomics and translatomics datasets. Our data shows that unlike RNA changes protein changes are less dynamic. Further increases in protein levels correlate with mRNA translation whereas declines in protein levels do not suggesting active protein degradation processes. Interestingly proteins from pure zygotic genes are present at fertilization challenging existing mRNA based gene classifications. As a proof of concept we utilized CRISPR Cas13d to target znf281b mRNA a gene whose protein significantly accumulates within the first two hpf demonstrating its crucial role in development. Consequently our protein profiling coupled with CRISPR Cas13d offers a new approach to unravel maternal mRNAs function during embryonic development. Overall design: Two biological replicate samples containing 50 zebrafish oocytes per stage were collected by pairing females in natural matings to ''purge'' mature eggs and used to establish an oogenesis time line. Fish were euthanized and ovaries harvested within 1–11 days post purging dpp. Stage I and II oocytes were collected at 1–2 dpp stage III oocytes germinal vesicle in central position at 4–7 dpp and stage IV oocytes germinal vesicle asymmetrically located at 8–10 dpp. Oocyte isolations were based on73 and conducted in isolation medium Leibovitz L 15 Sigma Aldrich #L5520 plus Collagenase I and II depending on the stages as follows. Stages I and II were isolated with 3 mg/mL Collagenase I Sigma Aldrich C0130 and 3 mg/mL Collagenase II Gibco 17101015. Stage III was isolated with 3 mg/mL Collagenase I Sigma Aldrich C0130. Stage IV was isolated by mechanical stripping with forceps and nee…   pubmed:39302832   Zebrafish Embryo 4hpf rep1 GSM8147859   source name:Whole embryo|tissue:Whole embryo|developmental stage:4hpf|genotype:AB TF and TLF|geo loc name:missing|collection date:missing Zebrafish Embryo 4hpf rep1 Raw reads from zebrafish oocytes stages I IV and embryos 0 2 4 and 6 hpf were demultiplexed into FASTQ format allowing up to one mismatch using Illumina bcl convert 3.10.5. Reads were aligned to UCSC genome danRer11 with STAR aligner version 2.7.3a using Ensembl 106 gene models. Counts were converted to TPM values using RSEM version v1.3.0 and all subsequent analysis was done using TPM. Assembly: danRer11 Ensembl 106 gene models Supplementary files format and content: comma separated files with TPMs from all replicates Whole embryo   Isolated oocytes were snap frozen and RNA was extracted with TRIzol Invitrogen #15596026 following manufacturer’s protocols. 0 2 4 and 6 hpf zebrafish embryo RNA was extracted with Direct zol RNA MicroPrep kit Zymo Research #R2062 following manufacturer’s protocols. Ribo dep Stranded RNA Seq Zebrafish embryos 0 2 4 and 6 hpf total RNA sequencing libraries were generated from 500ng of high quality total RNA as assessed using Bioanalyzer Agilent. Libraries were made according to the manufacturer’s directions for the TruSeq Stranded Total RNA Library Prep Gold Illumina #20020598 and TruSeq RNA Single Indexes Sets A and B Illumina #20020492 and #20020493. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer Agilent Technologies and Qubit Fluorometer Life Technologies. Libraries were pooled quantified and sequenced as 75bp single reads on a high output flow cell using the Illumina NextSeq500 instrument. Following sequencing Illumina Primary Analysis version RTA 2.11.3.0 and bcl convert 3.10.5 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish oocytes stages I IV total RNA Seq libraries were generated from 100 ng of high quality total RNA as assessed by a Bioanalyzer Agilent Technologies. Libraries were prepared according to manufacturer’s instructions using the TruSeq Stranded Total RNA Library Prep Gold Illumina Cat. No. 20020598 and TruSeq RNA Single Indexes; Sets A and B Illumina Cat. No. 20020492 and 20020493. Resulting short fragment libraries were checked for quality and quantity using a Bioanalyzer and a Qubit Fluorometer Life Technologies. Libraries were normalized pooled and sequenced on a NextSeq 500 instrument Illumina as 75 bp single end reads on a high output flowcell using NextSeq Control Software 2.2.0.4. Following sequencing Illumina Primary Analysis version NextSeq RTA 2.4.11 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files.   tissue:Whole embryo|developmental stage:4hpf|genotype:AB TF and TLF GSM8147859 GSM8147859: Zebrafish Embryo 4hpf rep1; Danio rerio; RNA Seq GSM8147859 r1 GSM8147859 1 Isolated oocytes were snap frozen and RNA was extracted with TRIzol Invitrogen #15596026 following manufacturer's protocols. 0 2 4 and 6 hpf zebrafish embryo RNA was extracted with Direct zol RNA MicroPrep kit Zymo Research #R2062 following manufacturer's protocols. Ribo dep Stranded RNA Seq Zebrafish embryos 0 2 4 and 6 hpf total RNA sequencing libraries were generated from 500ng of high quality total RNA as assessed using Bioanalyzer Agilent. Libraries were made according to the manufacturer's directions for the TruSeq Stranded Total RNA Library Prep Gold Illumina #20020598 and TruSeq RNA Single Indexes Sets A and B Illumina #20020492 and #20020493. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer Agilent Technologies and Qubit Fluorometer Life Technologies. Libraries were pooled quantified and sequenced as 75bp single reads on a high output flow cell using the Illumina NextSeq500 instrument. Following sequencing Illumina Primary Analysis version RTA 2.11.3.0 and bcl convert 3.10.5 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish oocytes stages I IV total RNA Seq libraries were generated from 100 ng of high quality total RNA as assessed by a Bioanalyzer Agilent Technologies. Libraries were prepared according to manufacturer's instructions using the TruSeq Stranded Total RNA Library Prep Gold Illumina Cat. No. 20020598 and TruSeq RNA Single Indexes; Sets A and B Illumina Cat. No. 20020492 and 20020493. Resulting short fragment libraries were checked for quality and quantity using a Bioanalyzer and a Qubit Fluorometer Life Technologies. Libraries were normalized pooled and sequenced on a NextSeq 500 instrument Illumina as 75 bp single end reads on a high output flowcell using NextSeq Control Software 2.2.0.4. Following sequencing Illumina Primary Analysis version NextSeq RTA 2.4.11 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files.   RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA NextSeq 500   SRP495323     s_4hpf_1.fastq fastq 1494269972.0 19661447.0 GSM8147859 r1 0:76 A:365052201;C:369717962;G:351357642;T:408073055;N:69112 76       365052201 369717962 351357642 408073055 69112 SRX23954962 SRS20755383 SRA1824599 Computational Biology, Stowers Institute for Medical Research Computational Biology, Stowers Institute for Medical Research                       B   usable mapping rate illumina nextseq unknown cdna_unspecified trueseq bulk bulk bulk   United States 2024-03-14 Blastula Embryo Whole Organism All anatomical structures
34032 34032 SRR31033342 SRX26418811 SRS22938447 SRP539223 PRJNA1174207 Rbm24 dictates mRNA recruitment for germ plasm assembly GSE279756 Transcriptome Analysis Ribonucleoprotein RNP granules are the most common membrane less biomolecular condensates. However the mechanisms underlying their assembly are largely unknown. The aggregation of germ plasm determines the fate of primordial germ cells PGCs and serves as a model for RNP granule assembly. Here we show that maternal RNA binding protein Rbm24a is the key factor governing specific sorting of mRNAs. Mechanistically Rbm24a complexes with Buc and interacts to dictate the specific grasp of germ plasm mRNAs into phase separated condensates. Germ plasm particles lacking Rbm24a and mRNAs fail to undergo kinesin dependent transport towards the cleavage furrows where small granules fuse into large aggregates. Therefore the loss of maternal Rbm24a causes a complete degradation of germ plasm and the disappearance of PGCs. These findings demonstrate that Rbm24a functions as a nucleating organizer of the germ plasm highlighting an emerging mechanism for RNA binding proteins in reading and recruiting RNA components into the phase separated protein scaffold. Overall design: To examine the global transcriptome alterations following the loss of maternal Rbm24a we conducted bulk RNA seq analysis on Mrbm24a and their sibling embyos at 4 cell sphere stage and 24 hpf.       Zebrafish Riboseq M sphere 5 GSM8579734   source name:whole embryo|tissue:whole embryo|Stage:sphere stage|cell type:embryonic cell|genotype:Mrbm24a|geo loc name:missing|collection date:missing Zebrafish Riboseq M sphere 5 Each library was amplified by a 11 cycle PCR and 150 bp paired end sequencing was subjected to Illumina HiSeq 2000 to obtain the raw data. Assembly: GRCz10 Supplementary files format and content: tab delimited text file includes raw counts for each Sample Supplementary files format and content: tab delimited text file includes FPKM values for each Sample whole embryo   Total RNAs were extracted from WT and 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 total RNA was used for following library preparation. The polyA mRNA isolation was performed using OligodT beads. The mRNA fragmentation was performed using divalent cations and high temperature. Priming was performed using Random Primers. First strand cDNA and the second strand cDNA were synthesized. The purified double stranded cDNA was then treated to repair both ends and add a dA tailing in one reaction followed by a T A ligation to add adaptors to both ends. Size selection of Adaptor ligated DNA was then performed using DNA Clean Beads. Each sample was then amplified by PCR using P5 and P7 primers and the PCR products were validated. Then libraries with different indexs were multiplexed and loaded on an Illumina HiSeq/ Illumina Novaseq/ MGI2000 instrument for sequencing using a 2x150 paired end PE configuration according to manufacturer’s instructions. Zebrafish embryos were maintained in 1/3x Ringer's at 28.5C tissue:whole embryo|Stage:sphere stage|cell type:embryonic cell|genotype:Mrbm24a GSM8579734 GSM8579734: Zebrafish Riboseq M sphere 5; Danio rerio; RNA Seq GSM8579734 r1 GSM8579734 1 Total RNAs were extracted from WT and 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 total RNA was used for following library preparation. The polyA mRNA isolation was performed using OligodT beads. The mRNA fragmentation was performed using divalent cations and high temperature. Priming was performed using Random Primers. First strand cDNA and the second strand cDNA were synthesized. The purified double stranded cDNA was then treated to repair both ends and add a dA tailing in one reaction followed by a T A ligation to add adaptors to both ends. Size selection of Adaptor ligated DNA was then performed using DNA Clean Beads. Each sample was then amplified by PCR using P5 and P7 primers and the PCR products were validated. Then libraries with different indexs were multiplexed and loaded on an Illumina HiSeq/ Illumina Novaseq/ MGI2000 instrument for sequencing using a 2x150 paired end PE configuration according to manufacturer's instructions.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 2000   SRP539223     M-sphere-5_L1_1.fq.gz M-sphere-5_L1_2.fq.gz fastq fastq 6386202900.0 21287343.0 GSM8579734 r1 0:150 1:150 A:1749323046;C:1447024706;G:1497348312;T:1692462495;N:44341 150 150     1749323046 1447024706 1497348312 1692462495 44341 SRX26418811 SRS22938447 SRA1993092 Shandong university Shandong university                       B B biological fallback assumption illumina hiseq_era unknown poly_a unknown bulk bulk bulk   China 2024-10-17 Blastula Embryo Whole Organism All anatomical structures
34033 34033 SRR31033343 SRX26418810 SRS22938444 SRP539223 PRJNA1174207 Rbm24 dictates mRNA recruitment for germ plasm assembly GSE279756 Transcriptome Analysis Ribonucleoprotein RNP granules are the most common membrane less biomolecular condensates. However the mechanisms underlying their assembly are largely unknown. The aggregation of germ plasm determines the fate of primordial germ cells PGCs and serves as a model for RNP granule assembly. Here we show that maternal RNA binding protein Rbm24a is the key factor governing specific sorting of mRNAs. Mechanistically Rbm24a complexes with Buc and interacts to dictate the specific grasp of germ plasm mRNAs into phase separated condensates. Germ plasm particles lacking Rbm24a and mRNAs fail to undergo kinesin dependent transport towards the cleavage furrows where small granules fuse into large aggregates. Therefore the loss of maternal Rbm24a causes a complete degradation of germ plasm and the disappearance of PGCs. These findings demonstrate that Rbm24a functions as a nucleating organizer of the germ plasm highlighting an emerging mechanism for RNA binding proteins in reading and recruiting RNA components into the phase separated protein scaffold. Overall design: To examine the global transcriptome alterations following the loss of maternal Rbm24a we conducted bulk RNA seq analysis on Mrbm24a and their sibling embyos at 4 cell sphere stage and 24 hpf.       Zebrafish Riboseq M sphere 4 GSM8579733   source name:whole embryo|tissue:whole embryo|Stage:sphere stage|cell type:embryonic cell|genotype:Mrbm24a|geo loc name:missing|collection date:missing Zebrafish Riboseq M sphere 4 Each library was amplified by a 11 cycle PCR and 150 bp paired end sequencing was subjected to Illumina HiSeq 2000 to obtain the raw data. Assembly: GRCz10 Supplementary files format and content: tab delimited text file includes raw counts for each Sample Supplementary files format and content: tab delimited text file includes FPKM values for each Sample whole embryo   Total RNAs were extracted from WT and 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 total RNA was used for following library preparation. The polyA mRNA isolation was performed using OligodT beads. The mRNA fragmentation was performed using divalent cations and high temperature. Priming was performed using Random Primers. First strand cDNA and the second strand cDNA were synthesized. The purified double stranded cDNA was then treated to repair both ends and add a dA tailing in one reaction followed by a T A ligation to add adaptors to both ends. Size selection of Adaptor ligated DNA was then performed using DNA Clean Beads. Each sample was then amplified by PCR using P5 and P7 primers and the PCR products were validated. Then libraries with different indexs were multiplexed and loaded on an Illumina HiSeq/ Illumina Novaseq/ MGI2000 instrument for sequencing using a 2x150 paired end PE configuration according to manufacturer’s instructions. Zebrafish embryos were maintained in 1/3x Ringer's at 28.5C tissue:whole embryo|Stage:sphere stage|cell type:embryonic cell|genotype:Mrbm24a GSM8579733 GSM8579733: Zebrafish Riboseq M sphere 4; Danio rerio; RNA Seq GSM8579733 r1 GSM8579733 1 Total RNAs were extracted from WT and 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 total RNA was used for following library preparation. The polyA mRNA isolation was performed using OligodT beads. The mRNA fragmentation was performed using divalent cations and high temperature. Priming was performed using Random Primers. First strand cDNA and the second strand cDNA were synthesized. The purified double stranded cDNA was then treated to repair both ends and add a dA tailing in one reaction followed by a T A ligation to add adaptors to both ends. Size selection of Adaptor ligated DNA was then performed using DNA Clean Beads. Each sample was then amplified by PCR using P5 and P7 primers and the PCR products were validated. Then libraries with different indexs were multiplexed and loaded on an Illumina HiSeq/ Illumina Novaseq/ MGI2000 instrument for sequencing using a 2x150 paired end PE configuration according to manufacturer's instructions.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 2000   SRP539223     M-sphere-4_L1_1.fq.gz M-sphere-4_L1_2.fq.gz fastq fastq 7329708000.0 24432360.0 GSM8579733 r1 0:150 1:150 A:1999459748;C:1667498786;G:1726069847;T:1936629080;N:50539 150 150     1999459748 1667498786 1726069847 1936629080 50539 SRX26418810 SRS22938444 SRA1993092 Shandong university Shandong university                       B B biological fallback assumption illumina hiseq_era unknown poly_a unknown bulk bulk bulk   China 2024-10-17 Blastula Embryo Whole Organism All anatomical structures
34034 34034 SRR31033344 SRX26418809 SRS22938445 SRP539223 PRJNA1174207 Rbm24 dictates mRNA recruitment for germ plasm assembly GSE279756 Transcriptome Analysis Ribonucleoprotein RNP granules are the most common membrane less biomolecular condensates. However the mechanisms underlying their assembly are largely unknown. The aggregation of germ plasm determines the fate of primordial germ cells PGCs and serves as a model for RNP granule assembly. Here we show that maternal RNA binding protein Rbm24a is the key factor governing specific sorting of mRNAs. Mechanistically Rbm24a complexes with Buc and interacts to dictate the specific grasp of germ plasm mRNAs into phase separated condensates. Germ plasm particles lacking Rbm24a and mRNAs fail to undergo kinesin dependent transport towards the cleavage furrows where small granules fuse into large aggregates. Therefore the loss of maternal Rbm24a causes a complete degradation of germ plasm and the disappearance of PGCs. These findings demonstrate that Rbm24a functions as a nucleating organizer of the germ plasm highlighting an emerging mechanism for RNA binding proteins in reading and recruiting RNA components into the phase separated protein scaffold. Overall design: To examine the global transcriptome alterations following the loss of maternal Rbm24a we conducted bulk RNA seq analysis on Mrbm24a and their sibling embyos at 4 cell sphere stage and 24 hpf.       Zebrafish Riboseq M sphere 2 GSM8579732   source name:whole embryo|tissue:whole embryo|Stage:sphere stage|cell type:embryonic cell|genotype:Mrbm24a|geo loc name:missing|collection date:missing Zebrafish Riboseq M sphere 2 Each library was amplified by a 11 cycle PCR and 150 bp paired end sequencing was subjected to Illumina HiSeq 2000 to obtain the raw data. Assembly: GRCz10 Supplementary files format and content: tab delimited text file includes raw counts for each Sample Supplementary files format and content: tab delimited text file includes FPKM values for each Sample whole embryo   Total RNAs were extracted from WT and 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 total RNA was used for following library preparation. The polyA mRNA isolation was performed using OligodT beads. The mRNA fragmentation was performed using divalent cations and high temperature. Priming was performed using Random Primers. First strand cDNA and the second strand cDNA were synthesized. The purified double stranded cDNA was then treated to repair both ends and add a dA tailing in one reaction followed by a T A ligation to add adaptors to both ends. Size selection of Adaptor ligated DNA was then performed using DNA Clean Beads. Each sample was then amplified by PCR using P5 and P7 primers and the PCR products were validated. Then libraries with different indexs were multiplexed and loaded on an Illumina HiSeq/ Illumina Novaseq/ MGI2000 instrument for sequencing using a 2x150 paired end PE configuration according to manufacturer’s instructions. Zebrafish embryos were maintained in 1/3x Ringer's at 28.5C tissue:whole embryo|Stage:sphere stage|cell type:embryonic cell|genotype:Mrbm24a GSM8579732 GSM8579732: Zebrafish Riboseq M sphere 2; Danio rerio; RNA Seq GSM8579732 r1 GSM8579732 1 Total RNAs were extracted from WT and 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 total RNA was used for following library preparation. The polyA mRNA isolation was performed using OligodT beads. The mRNA fragmentation was performed using divalent cations and high temperature. Priming was performed using Random Primers. First strand cDNA and the second strand cDNA were synthesized. The purified double stranded cDNA was then treated to repair both ends and add a dA tailing in one reaction followed by a T A ligation to add adaptors to both ends. Size selection of Adaptor ligated DNA was then performed using DNA Clean Beads. Each sample was then amplified by PCR using P5 and P7 primers and the PCR products were validated. Then libraries with different indexs were multiplexed and loaded on an Illumina HiSeq/ Illumina Novaseq/ MGI2000 instrument for sequencing using a 2x150 paired end PE configuration according to manufacturer's instructions.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 2000   SRP539223     M-sphere-2_L1_1.fq.gz M-sphere-2_L1_2.fq.gz fastq fastq 6614502300.0 22048341.0 GSM8579732 r1 0:150 1:150 A:1811784017;C:1496173021;G:1549885996;T:1756614738;N:44528 150 150     1811784017 1496173021 1549885996 1756614738 44528 SRX26418809 SRS22938445 SRA1993092 Shandong university Shandong university                       B B biological fallback assumption illumina hiseq_era unknown poly_a unknown bulk bulk bulk   China 2024-10-17 Blastula Embryo Whole Organism All anatomical structures
34035 34035 SRR31033352 SRX26418808 SRS22938439 SRP539223 PRJNA1174207 Rbm24 dictates mRNA recruitment for germ plasm assembly GSE279756 Transcriptome Analysis Ribonucleoprotein RNP granules are the most common membrane less biomolecular condensates. However the mechanisms underlying their assembly are largely unknown. The aggregation of germ plasm determines the fate of primordial germ cells PGCs and serves as a model for RNP granule assembly. Here we show that maternal RNA binding protein Rbm24a is the key factor governing specific sorting of mRNAs. Mechanistically Rbm24a complexes with Buc and interacts to dictate the specific grasp of germ plasm mRNAs into phase separated condensates. Germ plasm particles lacking Rbm24a and mRNAs fail to undergo kinesin dependent transport towards the cleavage furrows where small granules fuse into large aggregates. Therefore the loss of maternal Rbm24a causes a complete degradation of germ plasm and the disappearance of PGCs. These findings demonstrate that Rbm24a functions as a nucleating organizer of the germ plasm highlighting an emerging mechanism for RNA binding proteins in reading and recruiting RNA components into the phase separated protein scaffold. Overall design: To examine the global transcriptome alterations following the loss of maternal Rbm24a we conducted bulk RNA seq analysis on Mrbm24a and their sibling embyos at 4 cell sphere stage and 24 hpf.       Zebrafish Riboseq M sphere 1 GSM8579731   source name:whole embryo|tissue:whole embryo|Stage:sphere stage|cell type:embryonic cell|genotype:Mrbm24a|geo loc name:missing|collection date:missing Zebrafish Riboseq M sphere 1 Each library was amplified by a 11 cycle PCR and 150 bp paired end sequencing was subjected to Illumina HiSeq 2000 to obtain the raw data. Assembly: GRCz10 Supplementary files format and content: tab delimited text file includes raw counts for each Sample Supplementary files format and content: tab delimited text file includes FPKM values for each Sample whole embryo   Total RNAs were extracted from WT and 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 total RNA was used for following library preparation. The polyA mRNA isolation was performed using OligodT beads. The mRNA fragmentation was performed using divalent cations and high temperature. Priming was performed using Random Primers. First strand cDNA and the second strand cDNA were synthesized. The purified double stranded cDNA was then treated to repair both ends and add a dA tailing in one reaction followed by a T A ligation to add adaptors to both ends. Size selection of Adaptor ligated DNA was then performed using DNA Clean Beads. Each sample was then amplified by PCR using P5 and P7 primers and the PCR products were validated. Then libraries with different indexs were multiplexed and loaded on an Illumina HiSeq/ Illumina Novaseq/ MGI2000 instrument for sequencing using a 2x150 paired end PE configuration according to manufacturer’s instructions. Zebrafish embryos were maintained in 1/3x Ringer's at 28.5C tissue:whole embryo|Stage:sphere stage|cell type:embryonic cell|genotype:Mrbm24a GSM8579731 GSM8579731: Zebrafish Riboseq M sphere 1; Danio rerio; RNA Seq GSM8579731 r1 GSM8579731 1 Total RNAs were extracted from WT and 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 total RNA was used for following library preparation. The polyA mRNA isolation was performed using OligodT beads. The mRNA fragmentation was performed using divalent cations and high temperature. Priming was performed using Random Primers. First strand cDNA and the second strand cDNA were synthesized. The purified double stranded cDNA was then treated to repair both ends and add a dA tailing in one reaction followed by a T A ligation to add adaptors to both ends. Size selection of Adaptor ligated DNA was then performed using DNA Clean Beads. Each sample was then amplified by PCR using P5 and P7 primers and the PCR products were validated. Then libraries with different indexs were multiplexed and loaded on an Illumina HiSeq/ Illumina Novaseq/ MGI2000 instrument for sequencing using a 2x150 paired end PE configuration according to manufacturer's instructions.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 2000   SRP539223     M-sphere-1_L1_1.fq.gz M-sphere-1_L1_2.fq.gz fastq fastq 7080587100.0 23601957.0 GSM8579731 r1 0:150 1:150 A:1946013215;C:1594400814;G:1652448966;T:1887676214;N:47891 150 150     1946013215 1594400814 1652448966 1887676214 47891 SRX26418808 SRS22938439 SRA1993092 Shandong university Shandong university                       B B biological fallback assumption illumina hiseq_era unknown poly_a unknown bulk bulk bulk   China 2024-10-17 Blastula Embryo Whole Organism All anatomical structures
34044 34044 SRR31033360 SRX26418799 SRS22938436 SRP539223 PRJNA1174207 Rbm24 dictates mRNA recruitment for germ plasm assembly GSE279756 Transcriptome Analysis Ribonucleoprotein RNP granules are the most common membrane less biomolecular condensates. However the mechanisms underlying their assembly are largely unknown. The aggregation of germ plasm determines the fate of primordial germ cells PGCs and serves as a model for RNP granule assembly. Here we show that maternal RNA binding protein Rbm24a is the key factor governing specific sorting of mRNAs. Mechanistically Rbm24a complexes with Buc and interacts to dictate the specific grasp of germ plasm mRNAs into phase separated condensates. Germ plasm particles lacking Rbm24a and mRNAs fail to undergo kinesin dependent transport towards the cleavage furrows where small granules fuse into large aggregates. Therefore the loss of maternal Rbm24a causes a complete degradation of germ plasm and the disappearance of PGCs. These findings demonstrate that Rbm24a functions as a nucleating organizer of the germ plasm highlighting an emerging mechanism for RNA binding proteins in reading and recruiting RNA components into the phase separated protein scaffold. Overall design: To examine the global transcriptome alterations following the loss of maternal Rbm24a we conducted bulk RNA seq analysis on Mrbm24a and their sibling embyos at 4 cell sphere stage and 24 hpf.       Zebrafish RNAseq Sibling sphere 4 GSM8579722   source name:whole embryo|tissue:whole embryo|Stage:sphere stage|cell type:embryonic cell|genotype:wild type|geo loc name:missing|collection date:missing Zebrafish RNAseq Sibling sphere 4 Each library was amplified by a 11 cycle PCR and 150 bp paired end sequencing was subjected to Illumina HiSeq 2000 to obtain the raw data. Assembly: GRCz10 Supplementary files format and content: tab delimited text file includes raw counts for each Sample Supplementary files format and content: tab delimited text file includes FPKM values for each Sample whole embryo   Total RNAs were extracted from WT and 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 total RNA was used for following library preparation. The polyA mRNA isolation was performed using OligodT beads. The mRNA fragmentation was performed using divalent cations and high temperature. Priming was performed using Random Primers. First strand cDNA and the second strand cDNA were synthesized. The purified double stranded cDNA was then treated to repair both ends and add a dA tailing in one reaction followed by a T A ligation to add adaptors to both ends. Size selection of Adaptor ligated DNA was then performed using DNA Clean Beads. Each sample was then amplified by PCR using P5 and P7 primers and the PCR products were validated. Then libraries with different indexs were multiplexed and loaded on an Illumina HiSeq/ Illumina Novaseq/ MGI2000 instrument for sequencing using a 2x150 paired end PE configuration according to manufacturer’s instructions. Zebrafish embryos were maintained in 1/3x Ringer's at 28.5C tissue:whole embryo|Stage:sphere stage|cell type:embryonic cell|genotype:wild type GSM8579722 GSM8579722: Zebrafish RNAseq Sibling sphere 4; Danio rerio; RNA Seq GSM8579722 r1 GSM8579722 1 Total RNAs were extracted from WT and 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 total RNA was used for following library preparation. The polyA mRNA isolation was performed using OligodT beads. The mRNA fragmentation was performed using divalent cations and high temperature. Priming was performed using Random Primers. First strand cDNA and the second strand cDNA were synthesized. The purified double stranded cDNA was then treated to repair both ends and add a dA tailing in one reaction followed by a T A ligation to add adaptors to both ends. Size selection of Adaptor ligated DNA was then performed using DNA Clean Beads. Each sample was then amplified by PCR using P5 and P7 primers and the PCR products were validated. Then libraries with different indexs were multiplexed and loaded on an Illumina HiSeq/ Illumina Novaseq/ MGI2000 instrument for sequencing using a 2x150 paired end PE configuration according to manufacturer's instructions.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 2000   SRP539223     Sibling-sphere-4_L1_1.fq.gz Sibling-sphere-4_L1_2.fq.gz fastq fastq 7056333000.0 23521110.0 GSM8579722 r1 0:150 1:150 A:1940035016;C:1589047995;G:1648953235;T:1878249310;N:47444 150 150     1940035016 1589047995 1648953235 1878249310 47444 SRX26418799 SRS22938436 SRA1993092 Shandong university Shandong university                       B B biological fallback assumption illumina hiseq_era unknown poly_a unknown bulk bulk bulk   China 2024-10-17 Blastula Embryo Whole Organism All anatomical structures
34045 34045 SRR31033354 SRX26418798 SRS22938429 SRP539223 PRJNA1174207 Rbm24 dictates mRNA recruitment for germ plasm assembly GSE279756 Transcriptome Analysis Ribonucleoprotein RNP granules are the most common membrane less biomolecular condensates. However the mechanisms underlying their assembly are largely unknown. The aggregation of germ plasm determines the fate of primordial germ cells PGCs and serves as a model for RNP granule assembly. Here we show that maternal RNA binding protein Rbm24a is the key factor governing specific sorting of mRNAs. Mechanistically Rbm24a complexes with Buc and interacts to dictate the specific grasp of germ plasm mRNAs into phase separated condensates. Germ plasm particles lacking Rbm24a and mRNAs fail to undergo kinesin dependent transport towards the cleavage furrows where small granules fuse into large aggregates. Therefore the loss of maternal Rbm24a causes a complete degradation of germ plasm and the disappearance of PGCs. These findings demonstrate that Rbm24a functions as a nucleating organizer of the germ plasm highlighting an emerging mechanism for RNA binding proteins in reading and recruiting RNA components into the phase separated protein scaffold. Overall design: To examine the global transcriptome alterations following the loss of maternal Rbm24a we conducted bulk RNA seq analysis on Mrbm24a and their sibling embyos at 4 cell sphere stage and 24 hpf.       Zebrafish RNAseq Sibling sphere 3 GSM8579721   source name:whole embryo|tissue:whole embryo|Stage:sphere stage|cell type:embryonic cell|genotype:wild type|geo loc name:missing|collection date:missing Zebrafish RNAseq Sibling sphere 3 Each library was amplified by a 11 cycle PCR and 150 bp paired end sequencing was subjected to Illumina HiSeq 2000 to obtain the raw data. Assembly: GRCz10 Supplementary files format and content: tab delimited text file includes raw counts for each Sample Supplementary files format and content: tab delimited text file includes FPKM values for each Sample whole embryo   Total RNAs were extracted from WT and 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 total RNA was used for following library preparation. The polyA mRNA isolation was performed using OligodT beads. The mRNA fragmentation was performed using divalent cations and high temperature. Priming was performed using Random Primers. First strand cDNA and the second strand cDNA were synthesized. The purified double stranded cDNA was then treated to repair both ends and add a dA tailing in one reaction followed by a T A ligation to add adaptors to both ends. Size selection of Adaptor ligated DNA was then performed using DNA Clean Beads. Each sample was then amplified by PCR using P5 and P7 primers and the PCR products were validated. Then libraries with different indexs were multiplexed and loaded on an Illumina HiSeq/ Illumina Novaseq/ MGI2000 instrument for sequencing using a 2x150 paired end PE configuration according to manufacturer’s instructions. Zebrafish embryos were maintained in 1/3x Ringer's at 28.5C tissue:whole embryo|Stage:sphere stage|cell type:embryonic cell|genotype:wild type GSM8579721 GSM8579721: Zebrafish RNAseq Sibling sphere 3; Danio rerio; RNA Seq GSM8579721 r1 GSM8579721 1 Total RNAs were extracted from WT and 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 total RNA was used for following library preparation. The polyA mRNA isolation was performed using OligodT beads. The mRNA fragmentation was performed using divalent cations and high temperature. Priming was performed using Random Primers. First strand cDNA and the second strand cDNA were synthesized. The purified double stranded cDNA was then treated to repair both ends and add a dA tailing in one reaction followed by a T A ligation to add adaptors to both ends. Size selection of Adaptor ligated DNA was then performed using DNA Clean Beads. Each sample was then amplified by PCR using P5 and P7 primers and the PCR products were validated. Then libraries with different indexs were multiplexed and loaded on an Illumina HiSeq/ Illumina Novaseq/ MGI2000 instrument for sequencing using a 2x150 paired end PE configuration according to manufacturer's instructions.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 2000   SRP539223     Sibling-sphere-3_L1_1.fq.gz Sibling-sphere-3_L1_2.fq.gz fastq fastq 7102058100.0 23673527.0 GSM8579721 r1 0:150 1:150 A:1932660580;C:1619713195;G:1685329939;T:1864306893;N:47493 150 150     1932660580 1619713195 1685329939 1864306893 47493 SRX26418798 SRS22938429 SRA1993092 Shandong university Shandong university                       B B biological fallback assumption illumina hiseq_era unknown poly_a unknown bulk bulk bulk   China 2024-10-17 Blastula Embryo Whole Organism All anatomical structures
34046 34046 SRR31033359 SRX26418797 SRS22938433 SRP539223 PRJNA1174207 Rbm24 dictates mRNA recruitment for germ plasm assembly GSE279756 Transcriptome Analysis Ribonucleoprotein RNP granules are the most common membrane less biomolecular condensates. However the mechanisms underlying their assembly are largely unknown. The aggregation of germ plasm determines the fate of primordial germ cells PGCs and serves as a model for RNP granule assembly. Here we show that maternal RNA binding protein Rbm24a is the key factor governing specific sorting of mRNAs. Mechanistically Rbm24a complexes with Buc and interacts to dictate the specific grasp of germ plasm mRNAs into phase separated condensates. Germ plasm particles lacking Rbm24a and mRNAs fail to undergo kinesin dependent transport towards the cleavage furrows where small granules fuse into large aggregates. Therefore the loss of maternal Rbm24a causes a complete degradation of germ plasm and the disappearance of PGCs. These findings demonstrate that Rbm24a functions as a nucleating organizer of the germ plasm highlighting an emerging mechanism for RNA binding proteins in reading and recruiting RNA components into the phase separated protein scaffold. Overall design: To examine the global transcriptome alterations following the loss of maternal Rbm24a we conducted bulk RNA seq analysis on Mrbm24a and their sibling embyos at 4 cell sphere stage and 24 hpf.       Zebrafish RNAseq Sibling sphere 1 GSM8579720   source name:whole embryo|tissue:whole embryo|Stage:sphere stage|cell type:embryonic cell|genotype:wild type|geo loc name:missing|collection date:missing Zebrafish RNAseq Sibling sphere 1 Each library was amplified by a 11 cycle PCR and 150 bp paired end sequencing was subjected to Illumina HiSeq 2000 to obtain the raw data. Assembly: GRCz10 Supplementary files format and content: tab delimited text file includes raw counts for each Sample Supplementary files format and content: tab delimited text file includes FPKM values for each Sample whole embryo   Total RNAs were extracted from WT and 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 total RNA was used for following library preparation. The polyA mRNA isolation was performed using OligodT beads. The mRNA fragmentation was performed using divalent cations and high temperature. Priming was performed using Random Primers. First strand cDNA and the second strand cDNA were synthesized. The purified double stranded cDNA was then treated to repair both ends and add a dA tailing in one reaction followed by a T A ligation to add adaptors to both ends. Size selection of Adaptor ligated DNA was then performed using DNA Clean Beads. Each sample was then amplified by PCR using P5 and P7 primers and the PCR products were validated. Then libraries with different indexs were multiplexed and loaded on an Illumina HiSeq/ Illumina Novaseq/ MGI2000 instrument for sequencing using a 2x150 paired end PE configuration according to manufacturer’s instructions. Zebrafish embryos were maintained in 1/3x Ringer's at 28.5C tissue:whole embryo|Stage:sphere stage|cell type:embryonic cell|genotype:wild type GSM8579720 GSM8579720: Zebrafish RNAseq Sibling sphere 1; Danio rerio; RNA Seq GSM8579720 r1 GSM8579720 1 Total RNAs were extracted from WT and 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 total RNA was used for following library preparation. The polyA mRNA isolation was performed using OligodT beads. The mRNA fragmentation was performed using divalent cations and high temperature. Priming was performed using Random Primers. First strand cDNA and the second strand cDNA were synthesized. The purified double stranded cDNA was then treated to repair both ends and add a dA tailing in one reaction followed by a T A ligation to add adaptors to both ends. Size selection of Adaptor ligated DNA was then performed using DNA Clean Beads. Each sample was then amplified by PCR using P5 and P7 primers and the PCR products were validated. Then libraries with different indexs were multiplexed and loaded on an Illumina HiSeq/ Illumina Novaseq/ MGI2000 instrument for sequencing using a 2x150 paired end PE configuration according to manufacturer's instructions.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 2000   SRP539223     Sibling-sphere-1_L1_1.fq.gz Sibling-sphere-1_L1_2.fq.gz fastq fastq 6375563100.0 21251877.0 GSM8579720 r1 0:150 1:150 A:1748656292;C:1440631635;G:1494630558;T:1691601145;N:43470 150 150     1748656292 1440631635 1494630558 1691601145 43470 SRX26418797 SRS22938433 SRA1993092 Shandong university Shandong university                       B B biological fallback assumption illumina hiseq_era unknown poly_a unknown bulk bulk bulk   China 2024-10-17 Blastula Embryo Whole Organism All anatomical structures
34364 34364 SRR31695142 SRX27056958 SRS23518718 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d + g1 Chemically Synthesized si:dkey 93m18.4 rep4 GSM8678154   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:Knockdown Chemically Synthesized gRNA|geo loc name:missing|collection date:missing RfxCas13d + g1 Chemically Synthesized si:dkey 93m18.4 rep4 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:Knockdown Chemically Synthesized gRNA GSM8678154 GSM8678154: RfxCas13d + g1 Chemically Synthesized si:dkey 93m18.4 rep4; Danio rerio; RNA Seq GSM8678154 r1 GSM8678154 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_g1Syn_rep4_R1.fastq.gz T_pRfxCas13d_g1Syn_rep4_R2.fastq.gz fastq fastq 1869396000.0 12462640.0 GSM8678154 r1 0:75 1:75 A:497545631;C:434254877;G:436435676;T:501090064;N:69752 75 75     497545631 434254877 436435676 501090064 69752 SRX27056958 SRS23518718 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34365 34365 SRR31695143 SRX27056957 SRS23518717 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d + g1 Chemically Synthesized si:dkey 93m18.4 rep3 GSM8678153   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:Knockdown Chemically Synthesized gRNA|geo loc name:missing|collection date:missing RfxCas13d + g1 Chemically Synthesized si:dkey 93m18.4 rep3 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:Knockdown Chemically Synthesized gRNA GSM8678153 GSM8678153: RfxCas13d + g1 Chemically Synthesized si:dkey 93m18.4 rep3; Danio rerio; RNA Seq GSM8678153 r1 GSM8678153 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_g1Syn_rep3_R1.fastq.gz T_pRfxCas13d_g1Syn_rep3_R2.fastq.gz fastq fastq 1939758450.0 12931723.0 GSM8678153 r1 0:75 1:75 A:510928336;C:455303431;G:458245636;T:515206838;N:74209 75 75     510928336 455303431 458245636 515206838 74209 SRX27056957 SRS23518717 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34366 34366 SRR31695144 SRX27056956 SRS23518716 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d + g3 Chemically Synthesized si:dkey 93m18.4 rep4 GSM8678152   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:Knockdown Chemically Synthesized gRNA|geo loc name:missing|collection date:missing RfxCas13d + g3 Chemically Synthesized si:dkey 93m18.4 rep4 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:Knockdown Chemically Synthesized gRNA GSM8678152 GSM8678152: RfxCas13d + g3 Chemically Synthesized si:dkey 93m18.4 rep4; Danio rerio; RNA Seq GSM8678152 r1 GSM8678152 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_g3Syn_rep4_R1.fastq.gz T_pRfxCas13d_g3Syn_rep4_R2.fastq.gz fastq fastq 1372458150.0 9149721.0 GSM8678152 r1 0:75 1:75 A:363668106;C:319963774;G:322725620;T:366049498;N:51152 75 75     363668106 319963774 322725620 366049498 51152 SRX27056956 SRS23518716 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34367 34367 SRR31695145 SRX27056955 SRS23518715 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d + g3 Chemically Synthesized si:dkey 93m18.4 rep3 GSM8678151   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:Knockdown Chemically Synthesized gRNA|geo loc name:missing|collection date:missing RfxCas13d + g3 Chemically Synthesized si:dkey 93m18.4 rep3 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:Knockdown Chemically Synthesized gRNA GSM8678151 GSM8678151: RfxCas13d + g3 Chemically Synthesized si:dkey 93m18.4 rep3; Danio rerio; RNA Seq GSM8678151 r1 GSM8678151 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_g3Syn_rep3_R1.fastq.gz T_pRfxCas13d_g3Syn_rep3_R2.fastq.gz fastq fastq 1716718500.0 11444790.0 GSM8678151 r1 0:75 1:75 A:454308432;C:401470287;G:402968652;T:457908108;N:63021 75 75     454308432 401470287 402968652 457908108 63021 SRX27056955 SRS23518715 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34368 34368 SRR31695150 SRX27056954 SRS23518714 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d control rep6 GSM8678150   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:RfxCas13d control|geo loc name:missing|collection date:missing RfxCas13d control rep6 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:RfxCas13d control GSM8678150 GSM8678150: RfxCas13d control rep6; Danio rerio; RNA Seq GSM8678150 r1 GSM8678150 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_rep6_R1.fastq.gz T_pRfxCas13d_rep6_R2.fastq.gz fastq fastq 1820554200.0 12137028.0 GSM8678150 r1 0:75 1:75 A:480817156;C:426951021;G:428763810;T:483952927;N:69286 75 75     480817156 426951021 428763810 483952927 69286 SRX27056954 SRS23518714 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34369 34369 SRR31695146 SRX27056953 SRS23518713 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d + g1 Chemically Synthesized si:dkey 93m18.4 rep2 GSM8678149   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:Knockdown Chemically Synthesized gRNA|geo loc name:missing|collection date:missing RfxCas13d + g1 Chemically Synthesized si:dkey 93m18.4 rep2 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:Knockdown Chemically Synthesized gRNA GSM8678149 GSM8678149: RfxCas13d + g1 Chemically Synthesized si:dkey 93m18.4 rep2; Danio rerio; RNA Seq GSM8678149 r1 GSM8678149 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_g1Syn_rep2_R1.fastq.gz T_pRfxCas13d_g1Syn_rep2_R2.fastq.gz fastq fastq 1686506550.0 11243377.0 GSM8678149 r1 0:75 1:75 A:445306390;C:395515087;G:396951869;T:448671449;N:61755 75 75     445306390 395515087 396951869 448671449 61755 SRX27056953 SRS23518713 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34370 34370 SRR31695147 SRX27056952 SRS23518712 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d + g1 Chemically Synthesized si:dkey 93m18.4 rep1 GSM8678148   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:Knockdown Chemically Synthesized gRNA|geo loc name:missing|collection date:missing RfxCas13d + g1 Chemically Synthesized si:dkey 93m18.4 rep1 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:Knockdown Chemically Synthesized gRNA GSM8678148 GSM8678148: RfxCas13d + g1 Chemically Synthesized si:dkey 93m18.4 rep1; Danio rerio; RNA Seq GSM8678148 r1 GSM8678148 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_g1Syn_rep1_R1.fastq.gz T_pRfxCas13d_g1Syn_rep1_R2.fastq.gz fastq fastq 2099239050.0 13994927.0 GSM8678148 r1 0:75 1:75 A:555464950;C:490168575;G:492715156;T:560811934;N:78435 75 75     555464950 490168575 492715156 560811934 78435 SRX27056952 SRS23518712 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34371 34371 SRR31695148 SRX27056951 SRS23518711 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d + g3 Chemically Synthesized si:dkey 93m18.4 rep2 GSM8678147   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:Knockdown Chemically Synthesized gRNA|geo loc name:missing|collection date:missing RfxCas13d + g3 Chemically Synthesized si:dkey 93m18.4 rep2 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:Knockdown Chemically Synthesized gRNA GSM8678147 GSM8678147: RfxCas13d + g3 Chemically Synthesized si:dkey 93m18.4 rep2; Danio rerio; RNA Seq GSM8678147 r1 GSM8678147 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_g3Syn_rep2_R1.fastq.gz T_pRfxCas13d_g3Syn_rep2_R2.fastq.gz fastq fastq 1855616550.0 12370777.0 GSM8678147 r1 0:75 1:75 A:490735862;C:434070234;G:436325424;T:494415408;N:69622 75 75     490735862 434070234 436325424 494415408 69622 SRX27056951 SRS23518711 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34372 34372 SRR31695149 SRX27056950 SRS23518710 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d + g3 Chemically Synthesized si:dkey 93m18.4 rep1 GSM8678146   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:Knockdown Chemically Synthesized gRNA|geo loc name:missing|collection date:missing RfxCas13d + g3 Chemically Synthesized si:dkey 93m18.4 rep1 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:Knockdown Chemically Synthesized gRNA GSM8678146 GSM8678146: RfxCas13d + g3 Chemically Synthesized si:dkey 93m18.4 rep1; Danio rerio; RNA Seq GSM8678146 r1 GSM8678146 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_g3Syn_rep1_R1.fastq.gz T_pRfxCas13d_g3Syn_rep1_R2.fastq.gz fastq fastq 2029890750.0 13532605.0 GSM8678146 r1 0:75 1:75 A:536398630;C:474872197;G:478734292;T:539809587;N:76044 75 75     536398630 474872197 478734292 539809587 76044 SRX27056950 SRS23518710 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34373 34373 SRR31695155 SRX27056949 SRS23518709 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d control rep5 GSM8678145   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:RfxCas13d control|geo loc name:missing|collection date:missing RfxCas13d control rep5 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:RfxCas13d control GSM8678145 GSM8678145: RfxCas13d control rep5; Danio rerio; RNA Seq GSM8678145 r1 GSM8678145 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_rep5_R1.fastq.gz T_pRfxCas13d_rep5_R2.fastq.gz fastq fastq 1699800000.0 11332000.0 GSM8678145 r1 0:75 1:75 A:448834438;C:398037498;G:400242751;T:452619971;N:65342 75 75     448834438 398037498 400242751 452619971 65342 SRX27056949 SRS23518709 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34374 34374 SRR31695151 SRX27056948 SRS23518708 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d + g3 IVTed si:dkey 93m18.4 rep3 GSM8678144   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:Knockdown IVTed gRNA|geo loc name:missing|collection date:missing RfxCas13d + g3 IVTed si:dkey 93m18.4 rep3 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:Knockdown IVTed gRNA GSM8678144 GSM8678144: RfxCas13d + g3 IVTed si:dkey 93m18.4 rep3; Danio rerio; RNA Seq GSM8678144 r1 GSM8678144 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_g3_rep3_R1.fastq.gz T_pRfxCas13d_g3_rep3_R2.fastq.gz fastq fastq 1603289100.0 10688594.0 GSM8678144 r1 0:75 1:75 A:420349766;C:379529179;G:379756700;T:423592677;N:60778 75 75     420349766 379529179 379756700 423592677 60778 SRX27056948 SRS23518708 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34375 34375 SRR31695154 SRX27056947 SRS23518706 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d + g3 IVTed si:dkey 93m18.4 rep2 GSM8678143   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:Knockdown IVTed gRNA|geo loc name:missing|collection date:missing RfxCas13d + g3 IVTed si:dkey 93m18.4 rep2 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:Knockdown IVTed gRNA GSM8678143 GSM8678143: RfxCas13d + g3 IVTed si:dkey 93m18.4 rep2; Danio rerio; RNA Seq GSM8678143 r1 GSM8678143 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_g3_rep2_R1.fastq.gz T_pRfxCas13d_g3_rep2_R2.fastq.gz fastq fastq 1700840400.0 11338936.0 GSM8678143 r1 0:75 1:75 A:447219410;C:401375108;G:401866426;T:450317233;N:62223 75 75     447219410 401375108 401866426 450317233 62223 SRX27056947 SRS23518706 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34376 34376 SRR31695152 SRX27056946 SRS23518707 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d control rep4 GSM8678142   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:RfxCas13d control|geo loc name:missing|collection date:missing RfxCas13d control rep4 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:RfxCas13d control GSM8678142 GSM8678142: RfxCas13d control rep4; Danio rerio; RNA Seq GSM8678142 r1 GSM8678142 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_rep4_R1.fastq.gz T_pRfxCas13d_rep4_R2.fastq.gz fastq fastq 1390819050.0 9272127.0 GSM8678142 r1 0:75 1:75 A:365774543;C:327280225;G:328329658;T:369385513;N:49111 75 75     365774543 327280225 328329658 369385513 49111 SRX27056946 SRS23518707 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34377 34377 SRR31695153 SRX27056945 SRS23518705 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d + g3 IVTed si:dkey 93m18.4 rep1 GSM8678141   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:Knockdown IVTed gRNA|geo loc name:missing|collection date:missing RfxCas13d + g3 IVTed si:dkey 93m18.4 rep1 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:Knockdown IVTed gRNA GSM8678141 GSM8678141: RfxCas13d + g3 IVTed si:dkey 93m18.4 rep1; Danio rerio; RNA Seq GSM8678141 r1 GSM8678141 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_g3_rep1_R1.fastq.gz T_pRfxCas13d_g3_rep1_R2.fastq.gz fastq fastq 1979083650.0 13193891.0 GSM8678141 r1 0:75 1:75 A:521444446;C:467332323;G:468450230;T:521779923;N:76728 75 75     521444446 467332323 468450230 521779923 76728 SRX27056945 SRS23518705 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34378 34378 SRR31695156 SRX27056944 SRS23518704 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d control rep3 GSM8678140   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:RfxCas13d control|geo loc name:missing|collection date:missing RfxCas13d control rep3 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:RfxCas13d control GSM8678140 GSM8678140: RfxCas13d control rep3; Danio rerio; RNA Seq GSM8678140 r1 GSM8678140 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_rep3_R1.fastq.gz T_pRfxCas13d_rep3_R2.fastq.gz fastq fastq 1520356800.0 10135712.0 GSM8678140 r1 0:75 1:75 A:400447283;C:357034714;G:358393387;T:404426537;N:54879 75 75     400447283 357034714 358393387 404426537 54879 SRX27056944 SRS23518704 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34379 34379 SRR31695157 SRX27056943 SRS23518703 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d + g1 IVTed si:dkey 93m18.4 rep3 GSM8678139   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:Knockdown IVTed gRNA|geo loc name:missing|collection date:missing RfxCas13d + g1 IVTed si:dkey 93m18.4 rep3 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:Knockdown IVTed gRNA GSM8678139 GSM8678139: RfxCas13d + g1 IVTed si:dkey 93m18.4 rep3; Danio rerio; RNA Seq GSM8678139 r1 GSM8678139 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_g1_rep3_R1.fastq.gz T_pRfxCas13d_g1_rep3_R2.fastq.gz fastq fastq 1890203250.0 12601355.0 GSM8678139 r1 0:75 1:75 A:501821472;C:441868721;G:441485425;T:504956362;N:71270 75 75     501821472 441868721 441485425 504956362 71270 SRX27056943 SRS23518703 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34380 34380 SRR31695162 SRX27056942 SRS23518702 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d + g2 IVTed si:dkey 93m18.4 rep3 GSM8678138   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:Knockdown IVTed gRNA|geo loc name:missing|collection date:missing RfxCas13d + g2 IVTed si:dkey 93m18.4 rep3 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:Knockdown IVTed gRNA GSM8678138 GSM8678138: RfxCas13d + g2 IVTed si:dkey 93m18.4 rep3; Danio rerio; RNA Seq GSM8678138 r1 GSM8678138 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_g2_rep3_R1.fastq.gz T_pRfxCas13d_g2_rep3_R2.fastq.gz fastq fastq 1839509400.0 12263396.0 GSM8678138 r1 0:75 1:75 A:485779725;C:432231942;G:432933832;T:488492257;N:71644 75 75     485779725 432231942 432933832 488492257 71644 SRX27056942 SRS23518702 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34381 34381 SRR31695158 SRX27056941 SRS23518701 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d + g1 IVTed si:dkey 93m18.4 rep2 GSM8678137   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:Knockdown IVTed gRNA|geo loc name:missing|collection date:missing RfxCas13d + g1 IVTed si:dkey 93m18.4 rep2 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:Knockdown IVTed gRNA GSM8678137 GSM8678137: RfxCas13d + g1 IVTed si:dkey 93m18.4 rep2; Danio rerio; RNA Seq GSM8678137 r1 GSM8678137 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_g1_rep2_R1.fastq.gz T_pRfxCas13d_g1_rep2_R2.fastq.gz fastq fastq 1742339250.0 11615595.0 GSM8678137 r1 0:75 1:75 A:461346904;C:406656276;G:409937021;T:464333259;N:65790 75 75     461346904 406656276 409937021 464333259 65790 SRX27056941 SRS23518701 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34382 34382 SRR31695161 SRX27056940 SRS23518700 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d control rep2 GSM8678136   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:RfxCas13d control|geo loc name:missing|collection date:missing RfxCas13d control rep2 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:RfxCas13d control GSM8678136 GSM8678136: RfxCas13d control rep2; Danio rerio; RNA Seq GSM8678136 r1 GSM8678136 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_rep2_R1.fastq.gz T_pRfxCas13d_rep2_R2.fastq.gz fastq fastq 1800739800.0 12004932.0 GSM8678136 r1 0:75 1:75 A:477337887;C:421087001;G:421493244;T:480753457;N:68211 75 75     477337887 421087001 421493244 480753457 68211 SRX27056940 SRS23518700 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34383 34383 SRR31695159 SRX27056939 SRS23518699 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d + g2 IVTed si:dkey 93m18.4 rep2 GSM8678135   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:Knockdown IVTed gRNA|geo loc name:missing|collection date:missing RfxCas13d + g2 IVTed si:dkey 93m18.4 rep2 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:Knockdown IVTed gRNA GSM8678135 GSM8678135: RfxCas13d + g2 IVTed si:dkey 93m18.4 rep2; Danio rerio; RNA Seq GSM8678135 r1 GSM8678135 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_g2_rep2_R1.fastq.gz T_pRfxCas13d_g2_rep2_R2.fastq.gz fastq fastq 1752030300.0 11680202.0 GSM8678135 r1 0:75 1:75 A:462116558;C:411708555;G:412346341;T:465792629;N:66217 75 75     462116558 411708555 412346341 465792629 66217 SRX27056939 SRS23518699 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34384 34384 SRR31695160 SRX27056938 SRS23518698 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d + g1 IVTed si:dkey 93m18.4 rep1 GSM8678134   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:Knockdown IVTed gRNA|geo loc name:missing|collection date:missing RfxCas13d + g1 IVTed si:dkey 93m18.4 rep1 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:Knockdown IVTed gRNA GSM8678134 GSM8678134: RfxCas13d + g1 IVTed si:dkey 93m18.4 rep1; Danio rerio; RNA Seq GSM8678134 r1 GSM8678134 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_g1_rep1_R1.fastq.gz T_pRfxCas13d_g1_rep1_R2.fastq.gz fastq fastq 1724105250.0 11494035.0 GSM8678134 r1 0:75 1:75 A:456793309;C:403294318;G:403256373;T:460696014;N:65236 75 75     456793309 403294318 403256373 460696014 65236 SRX27056938 SRS23518698 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34385 34385 SRR31695163 SRX27056937 SRS23518697 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d + g2 IVTed si:dkey 93m18.4 rep1 GSM8678133   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:Knockdown IVTed gRNA|geo loc name:missing|collection date:missing RfxCas13d + g2 IVTed si:dkey 93m18.4 rep1 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:Knockdown IVTed gRNA GSM8678133 GSM8678133: RfxCas13d + g2 IVTed si:dkey 93m18.4 rep1; Danio rerio; RNA Seq GSM8678133 r1 GSM8678133 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_g2_rep1_R1.fastq.gz T_pRfxCas13d_g2_rep1_R2.fastq.gz fastq fastq 1903402350.0 12689349.0 GSM8678133 r1 0:75 1:75 A:501904806;C:447043548;G:448270395;T:506113883;N:69718 75 75     501904806 447043548 448270395 506113883 69718 SRX27056937 SRS23518697 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
34386 34386 SRR31695164 SRX27056936 SRS23518696 SRP551059 PRJNA1197382 Enhanced RNA targeting CRISPR Cas technology in zebrafish V GSE284120 Transcriptome Analysis Transient elimination of RNAs by CRISPR Cas13 systems has been widely used in basic and applied sciences over the last years. However the efficiency of the system can be enhanced in vivo and its application has generated controversy due to the recently described collateral activity in mammalian cells and mouse models. Here we have optimized the CRISPR RfxCas13d CasRx system for an optimized RNA targeting in vivo in zebrafish embryos by different and compatible approaches. These strategies include the use of chemically modified guide RNAs to increase and sustain mRNA knockdown an improved nuclear targeting and the evaluation of ex vivo computational models for predicting gRNA efficiency in vivo. Furthermore our study demonstrated that transient CRISPR RfxCas13d approaches can effectively deplete endogenous mRNAs in zebrafish embryos without xxx collateral effects except for targeting extremely abundant RNAs. For that we have implemented alternative RNA targeting CRISPR Cas systems with reduced or absent collateral activity in zebrafish embryos. Altogether these findings contribute to optimize CRISPR Cas technology for RNA targeting in zebrafish through transient approaches and assist the progress of potential implications for knockdown therapies in vivo. Overall design: To investigate the toxicity effect of IVTed gRNAs compared with chemically synthesized parent bioproject:PRJNA1128082 pubmed:40091120   RfxCas13d control rep1 GSM8678132   source name:whole embryo|tissue:whole embryo|age:4 hpf|treatment:RfxCas13d control|geo loc name:missing|collection date:missing RfxCas13d control rep1 Following sequencing Illumina Primary Analysis version NextSeq RTA 2.11.3.0 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files. Assembly: danrer11 Supplementary files format and content: tab delimited txt file includes raw counts for each Sample whole embryo   Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer’s instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   tissue:whole embryo|age:4 hpf|treatment:RfxCas13d control GSM8678132 GSM8678132: RfxCas13d control rep1; Danio rerio; RNA Seq GSM8678132 r1 GSM8678132 1 Total RNA then was extracted using Direct zol RNA Miniprep Kit #R2050 Zymo Research following manufacturer's instructions. Zebrafish embryos total RNA sequencing libraries were generated from 100 ng of high quality total RNA as assessed using the Bioanalyzer Agilent with the Watchmaker mRNA Library Prep Kit Watchmaker Cat. No. 7BK0001 at a 1/5th reaction volume. mRNA isolation and library construction were completed per protocol at the miniaturization scale. For Library amplification 2 µL of IDT indexed primers using xGen™ Stubby Adapter UDI Primers 96 reactions Cat No. 10005921 was added to 3 µL of library and 5 µL of Equinox Amplification Master Mix 2x. These libraries were then amplified for 12 cycles of PCR using an annealing temperature of 55°C according to step 10.2. 1.0x SPRISelect Beckman Coulter Cat. No. B23318 bead ratio was used during the Post Amplification Cleanup and post amplification libraries were eluted in 10 µL of Qiagen EB Qiagen Cat. No. 19086. Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer and the Qubit Flex Fluorometer Life Technologies. Equal molar libraries were pooled quantified and converted to process on the Element AVITI.   RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ELEMENT Element AVITI   SRP551059     T_pRfxCas13d_rep1_R1.fastq.gz T_pRfxCas13d_rep1_R2.fastq.gz fastq fastq 1980525450.0 13203503.0 GSM8678132 r1 0:75 1:75 A:522445746;C:464027491;G:467474918;T:526505284;N:72011 75 75     522445746 464027491 467474918 526505284 72011 SRX27056936 SRS23518696 SRA2032641 Stowers Institute for Medical Research Stowers Institute for Medical Research                       B B biological fallback assumption element element unknown poly_a unknown bulk bulk bulk   United States 2024-12-12 Blastula Embryo Whole Organism All anatomical structures
38348 38348 SRR1785168 SRX864139 SRS834998 SRP053099 PRJNA274362 Transcriptomics analysis of gene expression in 10 embryo development stages of Danio rerio GSE65556 Transcriptome Analysis RNA was isolated from Danio embryo cells using the TRIzol Invitrogen reagent by following the company manual. For all samples the RNA integrity was checked using an Agilent Bioanalyzer 2100. All samples showed a RIN RNA integrity number of higher than 9. Approximately 2.5 µg of total RNA was then used for library preparation using a TruSeq™ RNA Sample Prep Kit v2 Illumina San Diego CA USA according to the manufacturer's protocol.The libraries were sequenced using HiSeq2000 Illumina in single read mode creating reads with a length of 101 bp. Sequencing chemistry v2 Illumina was used and samples were multiplexed in two samples per lane. Overall design: Examination of gene expressive levels in embryo development of Danio       sphere cells rep1 GSM1600044   tissue:Danio embryo cells|developmental stage:embryo|cell type:sphere cells sphere cells rep1 Reads were aligned to theZv9 genome assembly using TopHat v2.0.9 For each sample RPKM was computed as the number of reads which map per kilobase of exon model per million mapped reads for each gene. Genome build: Zv9 Supplementary files format and content: RPKM for each gene in each development stage Danio embryo cells   RNA was isolated from Danio cells using the TRIzol Invitrogen reagent by following the company manual. For all samples the RNA integrity was checked using an Agilent Bioanalyzer 2100 Approximately 2.5 µg of total RNA was then used for library preparation using a TruSeq™ RNA Sample Prep Kit v2 Illumina San Diego CA USA according to the manufacturer’s protocol.The libraries were sequenced using HiSeq2000 Illumina in single read mode creating reads with a length of 50 bp.   developmental stage:embryo|cell type:sphere cells GSM1600044 GSM1600044: sphere cells rep1; Danio rerio; RNA Seq GSM1600044   1 RNA was isolated from Danio cells using the TRIzol Invitrogen reagent by following the company manual. For all samples the RNA integrity was checked using an Agilent Bioanalyzer 2100 Approximately 2.5 µg of total RNA was then used for library preparation using a TruSeq™ RNA Sample Prep Kit v2 Illumina San Diego CA USA according to the manufacturer’s protocol.The libraries were sequenced using HiSeq2000 Illumina in single read mode creating reads with a length of 50 bp. GEO Accession:GSM1600044 RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA Illumina HiSeq 2000   SRP053099     W10T_1.fastq.gz fastq 9786365912.0 96894712.0 GSM1600044 r1 0:101 A:2573654910;C:2329556867;G:2311713480;T:2571211477;N:229178 101       2573654910 2329556867 2311713480 2571211477 229178 SRX864139 SRS834998 SRA236187 GEO Beijing Institute of Genomics (BIG) of Chinese Academy of Sciences (CAS) 1 0.95572   0.05587   0.73657   0.48128   101   B   usable mapping rate illumina hiseq_era unknown cdna_unspecified trueseq bulk bulk bulk   China 2015-02-03 Blastula Embryo Embryo Imprecise All anatomical structures
38349 38349 SRR1785167 SRX864138 SRS834995 SRP053099 PRJNA274362 Transcriptomics analysis of gene expression in 10 embryo development stages of Danio rerio GSE65556 Transcriptome Analysis RNA was isolated from Danio embryo cells using the TRIzol Invitrogen reagent by following the company manual. For all samples the RNA integrity was checked using an Agilent Bioanalyzer 2100. All samples showed a RIN RNA integrity number of higher than 9. Approximately 2.5 µg of total RNA was then used for library preparation using a TruSeq™ RNA Sample Prep Kit v2 Illumina San Diego CA USA according to the manufacturer's protocol.The libraries were sequenced using HiSeq2000 Illumina in single read mode creating reads with a length of 101 bp. Sequencing chemistry v2 Illumina was used and samples were multiplexed in two samples per lane. Overall design: Examination of gene expressive levels in embryo development of Danio       1000 cell rep1 GSM1600043   tissue:Danio embryo cells|developmental stage:embryo|cell type:1000 cells 1000 cell rep1 Reads were aligned to theZv9 genome assembly using TopHat v2.0.9 For each sample RPKM was computed as the number of reads which map per kilobase of exon model per million mapped reads for each gene. Genome build: Zv9 Supplementary files format and content: RPKM for each gene in each development stage Danio embryo cells   RNA was isolated from Danio cells using the TRIzol Invitrogen reagent by following the company manual. For all samples the RNA integrity was checked using an Agilent Bioanalyzer 2100 Approximately 2.5 µg of total RNA was then used for library preparation using a TruSeq™ RNA Sample Prep Kit v2 Illumina San Diego CA USA according to the manufacturer’s protocol.The libraries were sequenced using HiSeq2000 Illumina in single read mode creating reads with a length of 50 bp.   developmental stage:embryo|cell type:1000 cells GSM1600043 GSM1600043: 1000 cell rep1; Danio rerio; RNA Seq GSM1600043   1 RNA was isolated from Danio cells using the TRIzol Invitrogen reagent by following the company manual. For all samples the RNA integrity was checked using an Agilent Bioanalyzer 2100 Approximately 2.5 µg of total RNA was then used for library preparation using a TruSeq™ RNA Sample Prep Kit v2 Illumina San Diego CA USA according to the manufacturer’s protocol.The libraries were sequenced using HiSeq2000 Illumina in single read mode creating reads with a length of 50 bp. GEO Accession:GSM1600043 RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA Illumina HiSeq 2000   SRP053099     W9T_1.fastq.gz fastq 8394219282.0 83111082.0 GSM1600043 r1 0:101 A:2195214424;C:2012486651;G:1991514912;T:2194810285;N:193010 101       2195214424 2012486651 1991514912 2194810285 193010 SRX864138 SRS834995 SRA236187 GEO Beijing Institute of Genomics (BIG) of Chinese Academy of Sciences (CAS) 1 0.95627   0.02474   0.75761   0.47804   101   B   usable mapping rate illumina hiseq_era unknown cdna_unspecified trueseq bulk bulk bulk   China 2015-02-03 Blastula Embryo Embryo Imprecise All anatomical structures
38350 38350 SRR1785166 SRX864137 SRS834994 SRP053099 PRJNA274362 Transcriptomics analysis of gene expression in 10 embryo development stages of Danio rerio GSE65556 Transcriptome Analysis RNA was isolated from Danio embryo cells using the TRIzol Invitrogen reagent by following the company manual. For all samples the RNA integrity was checked using an Agilent Bioanalyzer 2100. All samples showed a RIN RNA integrity number of higher than 9. Approximately 2.5 µg of total RNA was then used for library preparation using a TruSeq™ RNA Sample Prep Kit v2 Illumina San Diego CA USA according to the manufacturer's protocol.The libraries were sequenced using HiSeq2000 Illumina in single read mode creating reads with a length of 101 bp. Sequencing chemistry v2 Illumina was used and samples were multiplexed in two samples per lane. Overall design: Examination of gene expressive levels in embryo development of Danio       256 cell rep1 GSM1600042   tissue:Danio embryo cells|developmental stage:embryo|cell type:256 cell 256 cell rep1 Reads were aligned to theZv9 genome assembly using TopHat v2.0.9 For each sample RPKM was computed as the number of reads which map per kilobase of exon model per million mapped reads for each gene. Genome build: Zv9 Supplementary files format and content: RPKM for each gene in each development stage Danio embryo cells   RNA was isolated from Danio cells using the TRIzol Invitrogen reagent by following the company manual. For all samples the RNA integrity was checked using an Agilent Bioanalyzer 2100 Approximately 2.5 µg of total RNA was then used for library preparation using a TruSeq™ RNA Sample Prep Kit v2 Illumina San Diego CA USA according to the manufacturer’s protocol.The libraries were sequenced using HiSeq2000 Illumina in single read mode creating reads with a length of 50 bp.   developmental stage:embryo|cell type:256 cell GSM1600042 GSM1600042: 256 cell rep1; Danio rerio; RNA Seq GSM1600042   1 RNA was isolated from Danio cells using the TRIzol Invitrogen reagent by following the company manual. For all samples the RNA integrity was checked using an Agilent Bioanalyzer 2100 Approximately 2.5 µg of total RNA was then used for library preparation using a TruSeq™ RNA Sample Prep Kit v2 Illumina San Diego CA USA according to the manufacturer’s protocol.The libraries were sequenced using HiSeq2000 Illumina in single read mode creating reads with a length of 50 bp. GEO Accession:GSM1600042 RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA Illumina HiSeq 2000   SRP053099     W8T_1.fastq.gz fastq 7155728901.0 70848801.0 GSM1600042 r1 0:101 A:1868886025;C:1718971317;G:1698634177;T:1869072256;N:165126 101       1868886025 1718971317 1698634177 1869072256 165126 SRX864137 SRS834994 SRA236187 GEO Beijing Institute of Genomics (BIG) of Chinese Academy of Sciences (CAS) 1 0.95828   0.02188   0.76307   0.47861   101   B   usable mapping rate illumina hiseq_era unknown cdna_unspecified trueseq bulk bulk bulk   China 2015-02-03 Blastula Embryo Embryo Imprecise All anatomical structures
38351 38351 SRR1785165 SRX864136 SRS835002 SRP053099 PRJNA274362 Transcriptomics analysis of gene expression in 10 embryo development stages of Danio rerio GSE65556 Transcriptome Analysis RNA was isolated from Danio embryo cells using the TRIzol Invitrogen reagent by following the company manual. For all samples the RNA integrity was checked using an Agilent Bioanalyzer 2100. All samples showed a RIN RNA integrity number of higher than 9. Approximately 2.5 µg of total RNA was then used for library preparation using a TruSeq™ RNA Sample Prep Kit v2 Illumina San Diego CA USA according to the manufacturer's protocol.The libraries were sequenced using HiSeq2000 Illumina in single read mode creating reads with a length of 101 bp. Sequencing chemistry v2 Illumina was used and samples were multiplexed in two samples per lane. Overall design: Examination of gene expressive levels in embryo development of Danio       128 cell rep1 GSM1600041   tissue:Danio embryo cells|developmental stage:embryo|cell type:128 cell 128 cell rep1 Reads were aligned to theZv9 genome assembly using TopHat v2.0.9 For each sample RPKM was computed as the number of reads which map per kilobase of exon model per million mapped reads for each gene. Genome build: Zv9 Supplementary files format and content: RPKM for each gene in each development stage Danio embryo cells   RNA was isolated from Danio cells using the TRIzol Invitrogen reagent by following the company manual. For all samples the RNA integrity was checked using an Agilent Bioanalyzer 2100 Approximately 2.5 µg of total RNA was then used for library preparation using a TruSeq™ RNA Sample Prep Kit v2 Illumina San Diego CA USA according to the manufacturer’s protocol.The libraries were sequenced using HiSeq2000 Illumina in single read mode creating reads with a length of 50 bp.   developmental stage:embryo|cell type:128 cell GSM1600041 GSM1600041: 128 cell rep1; Danio rerio; RNA Seq GSM1600041   1 RNA was isolated from Danio cells using the TRIzol Invitrogen reagent by following the company manual. For all samples the RNA integrity was checked using an Agilent Bioanalyzer 2100 Approximately 2.5 µg of total RNA was then used for library preparation using a TruSeq™ RNA Sample Prep Kit v2 Illumina San Diego CA USA according to the manufacturer’s protocol.The libraries were sequenced using HiSeq2000 Illumina in single read mode creating reads with a length of 50 bp. GEO Accession:GSM1600041 RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA Illumina HiSeq 2000   SRP053099     W7T_1.fastq.gz fastq 11606209465.0 114912965.0 GSM1600041 r1 0:101 A:2959737203;C:2867533898;G:2855568487;T:2922947397;N:422480 101       2959737203 2867533898 2855568487 2922947397 422480 SRX864136 SRS835002 SRA236187 GEO Beijing Institute of Genomics (BIG) of Chinese Academy of Sciences (CAS) 1 0.95887   0.01798   0.76824   0.47695   101   B   usable mapping rate illumina hiseq_era unknown cdna_unspecified trueseq bulk bulk bulk   China 2015-02-03 Blastula Embryo Embryo Imprecise All anatomical structures
47617 47617 SRR6784691 SRX3744229 SRS3000697 SRP133517 PRJNA436017 Maternal mRNA degradation by TUT4 and TUT7 in the zebrafish maternal to zygotic transition GSE111152 Transcriptome Analysis During the maternal to zygotic transition MZT maternal RNAs are actively degraded and replaced by newly synthesized zygotic transcripts in a highly coordinated manner. However it remains largely unknown how maternal mRNA decay is triggered in early vertebrate embryos. Here through genome wide profiling of RNA abundance and three prime modification we show that uridylation is induced at the onset of maternal mRNA clearance. The temporal control of uridylation is conserved in vertebrates. When the homologs of terminal uridylyltransferases TUT4 and TUT7 TUT4/7 are depleted in zebrafish and Xenopus maternal mRNA clearance is significantly delayed leading to developmental defects during gastrulation. Short tailed mRNAs are selectively uridylated by TUT4/7 with the highly uridylated transcripts degraded faster during the MZT than those with unmodified polyA tails. Our study demonstrates that uridylation plays a crucial role in timely mRNA degradation thereby allowing the progression of early development. Overall design: Examination of zebrafish embryos injected with control or TUT4/7 morpholinos at 1 cell stage. RNA seq libraries were prepared at different time points across the MZT period. Three experimental sets rs1a rs2b and rs2 were conducted independently.   pubmed:29625039   Zebrafish 5 hpf whole embryo injected with TUT47MO Exp. ID: rs2 GSM3024083   tissue:Zebrafish whole embryo|hpf translation blocking TUT7 translation blocking|strain:AB Zebrafish 5 hpf whole embryo injected with TUT47MO Exp. ID: rs2 The sequences were basecalled by Illumina RTA embedded in NextSeq Control Software 1.3.2 by the standard workflow. Sequenced reads from RNA seq data were mapped to the GRCz10 genome with RefSeq release 104 annotations above using STAR 2.5.3a via RSEM 2.1.31. The gene level quantifications were performed using RSEM with the default options for strand specific and paired end RNA seq. The "rs1a" set of our libraries was between sample normalized using RUV seq 1.10.0. The standard workflow suggested by RUV seq did not work due to the global changes of maternal transcripts following the progression of embryogenesis. Instead we first chose a subset of spike in RNAs that have ≥ 256 “expected counts” of reads in at least 90% of samples. Then the counts were normalized by the RUVg algorithm k = 1. The geometric means of the normalized counts were used as size factors for the scale normalization of all other RNAs. The ERCC spike in reads in our second RNA seq dataset were too unreliable to be used as references. Even the raw read counts mappable to the reference spike in sequences were significantly out of the linear correlations between any two samples. As a workaround we picked the internal controls from the first set to find genes that express more than 500 normalized reads with ≤ 0.2 of the log2 largest difference between two samples. Fourteen genes satisfied the criteria to become a set of internal controls. The internal reference genes were used in place of spike in RNAs for processing counts by the modified indirect RUVg approach. Genome build: GRCz10 Supplementary files format and content: comma separated text file include RUVg normalized rs1a and rs1b or TPM normalized rs2 values for each sample. Zebrafish whole embryo Approximately 1 nl of the solution containing TUT4/7 or control morpholinos was injected into wild type zebrafish embryos at xxx cell stage. For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. Zebrafish embryos were obtained by the natural mating of wild type AB strain and grown in embryo medium at 28.5˚C. Embryos were staged according to standard morphological criteria. hpf translation blocking TUT7 translation blocking|strain:AB GSM3024083 GSM3024083: Zebrafish 5 hpf whole embryo injected with TUT47MO Exp. ID: rs2; Danio rerio; RNA Seq GSM3024083   1 For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. GEO Accession:GSM3024083 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP133517     rs2-TUT47MO-5hpf_R1.fastq.gz rs2-TUT47MO-5hpf_R2.fastq.gz fastq fastq 5956989896.0 39190723.0 GSM3024083 r1 0:76 1:76 A:1416702080;C:1543800970;G:1539813949;T:1454939015;N:1733882 76 76     1416702080 1543800970 1539813949 1454939015 1733882 SRX3744229 SRS3000697 SRA662359 GEO Narry Kim Lab, Center for RNA Research, Institute for Basic Science 2 0.9294 0.93064 0.16382 0.16164 0.75197 0.7529 0.6297 0.62272 76 76 B B biological fallback assumption illumina nextseq unknown poly_a ribozero bulk bulk bulk   South Korea 2018-02-26 Blastula Embryo Whole Organism All anatomical structures
47618 47618 SRR6784690 SRX3744228 SRS3000696 SRP133517 PRJNA436017 Maternal mRNA degradation by TUT4 and TUT7 in the zebrafish maternal to zygotic transition GSE111152 Transcriptome Analysis During the maternal to zygotic transition MZT maternal RNAs are actively degraded and replaced by newly synthesized zygotic transcripts in a highly coordinated manner. However it remains largely unknown how maternal mRNA decay is triggered in early vertebrate embryos. Here through genome wide profiling of RNA abundance and three prime modification we show that uridylation is induced at the onset of maternal mRNA clearance. The temporal control of uridylation is conserved in vertebrates. When the homologs of terminal uridylyltransferases TUT4 and TUT7 TUT4/7 are depleted in zebrafish and Xenopus maternal mRNA clearance is significantly delayed leading to developmental defects during gastrulation. Short tailed mRNAs are selectively uridylated by TUT4/7 with the highly uridylated transcripts degraded faster during the MZT than those with unmodified polyA tails. Our study demonstrates that uridylation plays a crucial role in timely mRNA degradation thereby allowing the progression of early development. Overall design: Examination of zebrafish embryos injected with control or TUT4/7 morpholinos at 1 cell stage. RNA seq libraries were prepared at different time points across the MZT period. Three experimental sets rs1a rs2b and rs2 were conducted independently.   pubmed:29625039   Zebrafish 3 hpf whole embryo injected with TUT47MO Exp. ID: rs2 GSM3024082   tissue:Zebrafish whole embryo|hpf translation blocking TUT7 translation blocking|strain:AB Zebrafish 3 hpf whole embryo injected with TUT47MO Exp. ID: rs2 The sequences were basecalled by Illumina RTA embedded in NextSeq Control Software 1.3.2 by the standard workflow. Sequenced reads from RNA seq data were mapped to the GRCz10 genome with RefSeq release 104 annotations above using STAR 2.5.3a via RSEM 2.1.31. The gene level quantifications were performed using RSEM with the default options for strand specific and paired end RNA seq. The "rs1a" set of our libraries was between sample normalized using RUV seq 1.10.0. The standard workflow suggested by RUV seq did not work due to the global changes of maternal transcripts following the progression of embryogenesis. Instead we first chose a subset of spike in RNAs that have ≥ 256 “expected counts” of reads in at least 90% of samples. Then the counts were normalized by the RUVg algorithm k = 1. The geometric means of the normalized counts were used as size factors for the scale normalization of all other RNAs. The ERCC spike in reads in our second RNA seq dataset were too unreliable to be used as references. Even the raw read counts mappable to the reference spike in sequences were significantly out of the linear correlations between any two samples. As a workaround we picked the internal controls from the first set to find genes that express more than 500 normalized reads with ≤ 0.2 of the log2 largest difference between two samples. Fourteen genes satisfied the criteria to become a set of internal controls. The internal reference genes were used in place of spike in RNAs for processing counts by the modified indirect RUVg approach. Genome build: GRCz10 Supplementary files format and content: comma separated text file include RUVg normalized rs1a and rs1b or TPM normalized rs2 values for each sample. Zebrafish whole embryo Approximately 1 nl of the solution containing TUT4/7 or control morpholinos was injected into wild type zebrafish embryos at xxx cell stage. For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. Zebrafish embryos were obtained by the natural mating of wild type AB strain and grown in embryo medium at 28.5˚C. Embryos were staged according to standard morphological criteria. hpf translation blocking TUT7 translation blocking|strain:AB GSM3024082 GSM3024082: Zebrafish 3 hpf whole embryo injected with TUT47MO Exp. ID: rs2; Danio rerio; RNA Seq GSM3024082   1 For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. GEO Accession:GSM3024082 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP133517     rs2-TUT47MO-3hpf_R1.fastq.gz rs2-TUT47MO-3hpf_R2.fastq.gz fastq fastq 5793171744.0 38112972.0 GSM3024082 r1 0:76 1:76 A:1406452628;C:1469719832;G:1468022600;T:1447279036;N:1697648 76 76     1406452628 1469719832 1468022600 1447279036 1697648 SRX3744228 SRS3000696 SRA662359 GEO Narry Kim Lab, Center for RNA Research, Institute for Basic Science 2 0.95626 0.95636 0.07094 0.0679 0.76098 0.76219 0.59322 0.59334 76 76 B B biological fallback assumption illumina nextseq unknown poly_a ribozero bulk bulk bulk   South Korea 2018-02-26 Blastula Embryo Whole Organism All anatomical structures
47619 47619 SRR6784689 SRX3744227 SRS3000695 SRP133517 PRJNA436017 Maternal mRNA degradation by TUT4 and TUT7 in the zebrafish maternal to zygotic transition GSE111152 Transcriptome Analysis During the maternal to zygotic transition MZT maternal RNAs are actively degraded and replaced by newly synthesized zygotic transcripts in a highly coordinated manner. However it remains largely unknown how maternal mRNA decay is triggered in early vertebrate embryos. Here through genome wide profiling of RNA abundance and three prime modification we show that uridylation is induced at the onset of maternal mRNA clearance. The temporal control of uridylation is conserved in vertebrates. When the homologs of terminal uridylyltransferases TUT4 and TUT7 TUT4/7 are depleted in zebrafish and Xenopus maternal mRNA clearance is significantly delayed leading to developmental defects during gastrulation. Short tailed mRNAs are selectively uridylated by TUT4/7 with the highly uridylated transcripts degraded faster during the MZT than those with unmodified polyA tails. Our study demonstrates that uridylation plays a crucial role in timely mRNA degradation thereby allowing the progression of early development. Overall design: Examination of zebrafish embryos injected with control or TUT4/7 morpholinos at 1 cell stage. RNA seq libraries were prepared at different time points across the MZT period. Three experimental sets rs1a rs2b and rs2 were conducted independently.   pubmed:29625039   Zebrafish 5 hpf whole embryo injected with ConMO Exp. ID: rs2 GSM3024081   tissue:Zebrafish whole embryo|hpf Zebrafish 5 hpf whole embryo injected with ConMO Exp. ID: rs2 The sequences were basecalled by Illumina RTA embedded in NextSeq Control Software 1.3.2 by the standard workflow. Sequenced reads from RNA seq data were mapped to the GRCz10 genome with RefSeq release 104 annotations above using STAR 2.5.3a via RSEM 2.1.31. The gene level quantifications were performed using RSEM with the default options for strand specific and paired end RNA seq. The "rs1a" set of our libraries was between sample normalized using RUV seq 1.10.0. The standard workflow suggested by RUV seq did not work due to the global changes of maternal transcripts following the progression of embryogenesis. Instead we first chose a subset of spike in RNAs that have ≥ 256 “expected counts” of reads in at least 90% of samples. Then the counts were normalized by the RUVg algorithm k = 1. The geometric means of the normalized counts were used as size factors for the scale normalization of all other RNAs. The ERCC spike in reads in our second RNA seq dataset were too unreliable to be used as references. Even the raw read counts mappable to the reference spike in sequences were significantly out of the linear correlations between any two samples. As a workaround we picked the internal controls from the first set to find genes that express more than 500 normalized reads with ≤ 0.2 of the log2 largest difference between two samples. Fourteen genes satisfied the criteria to become a set of internal controls. The internal reference genes were used in place of spike in RNAs for processing counts by the modified indirect RUVg approach. Genome build: GRCz10 Supplementary files format and content: comma separated text file include RUVg normalized rs1a and rs1b or TPM normalized rs2 values for each sample. Zebrafish whole embryo Approximately 1 nl of the solution containing TUT4/7 or control morpholinos was injected into wild type zebrafish embryos at xxx cell stage. For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. Zebrafish embryos were obtained by the natural mating of wild type AB strain and grown in embryo medium at 28.5˚C. Embryos were staged according to standard morphological criteria. hpf GSM3024081 GSM3024081: Zebrafish 5 hpf whole embryo injected with ConMO Exp. ID: rs2; Danio rerio; RNA Seq GSM3024081   1 For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. GEO Accession:GSM3024081 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP133517     rs2-ConMO-5hpf_R1.fastq.gz rs2-ConMO-5hpf_R2.fastq.gz fastq fastq 6789240664.0 44666057.0 GSM3024081 r1 0:76 1:76 A:1692447747;C:1683205250;G:1674097638;T:1737501588;N:1988441 76 76     1692447747 1683205250 1674097638 1737501588 1988441 SRX3744227 SRS3000695 SRA662359 GEO Narry Kim Lab, Center for RNA Research, Institute for Basic Science 2 0.93365 0.93288 0.18031 0.17541 0.75601 0.75801 0.51533 0.59789 76 76 B B biological fallback assumption illumina nextseq unknown poly_a ribozero bulk bulk bulk   South Korea 2018-02-26 Blastula Embryo Whole Organism All anatomical structures
47620 47620 SRR6784688 SRX3744226 SRS3000694 SRP133517 PRJNA436017 Maternal mRNA degradation by TUT4 and TUT7 in the zebrafish maternal to zygotic transition GSE111152 Transcriptome Analysis During the maternal to zygotic transition MZT maternal RNAs are actively degraded and replaced by newly synthesized zygotic transcripts in a highly coordinated manner. However it remains largely unknown how maternal mRNA decay is triggered in early vertebrate embryos. Here through genome wide profiling of RNA abundance and three prime modification we show that uridylation is induced at the onset of maternal mRNA clearance. The temporal control of uridylation is conserved in vertebrates. When the homologs of terminal uridylyltransferases TUT4 and TUT7 TUT4/7 are depleted in zebrafish and Xenopus maternal mRNA clearance is significantly delayed leading to developmental defects during gastrulation. Short tailed mRNAs are selectively uridylated by TUT4/7 with the highly uridylated transcripts degraded faster during the MZT than those with unmodified polyA tails. Our study demonstrates that uridylation plays a crucial role in timely mRNA degradation thereby allowing the progression of early development. Overall design: Examination of zebrafish embryos injected with control or TUT4/7 morpholinos at 1 cell stage. RNA seq libraries were prepared at different time points across the MZT period. Three experimental sets rs1a rs2b and rs2 were conducted independently.   pubmed:29625039   Zebrafish 3 hpf whole embryo injected with ConMO Exp. ID: rs2 GSM3024080   tissue:Zebrafish whole embryo|hpf Zebrafish 3 hpf whole embryo injected with ConMO Exp. ID: rs2 The sequences were basecalled by Illumina RTA embedded in NextSeq Control Software 1.3.2 by the standard workflow. Sequenced reads from RNA seq data were mapped to the GRCz10 genome with RefSeq release 104 annotations above using STAR 2.5.3a via RSEM 2.1.31. The gene level quantifications were performed using RSEM with the default options for strand specific and paired end RNA seq. The "rs1a" set of our libraries was between sample normalized using RUV seq 1.10.0. The standard workflow suggested by RUV seq did not work due to the global changes of maternal transcripts following the progression of embryogenesis. Instead we first chose a subset of spike in RNAs that have ≥ 256 “expected counts” of reads in at least 90% of samples. Then the counts were normalized by the RUVg algorithm k = 1. The geometric means of the normalized counts were used as size factors for the scale normalization of all other RNAs. The ERCC spike in reads in our second RNA seq dataset were too unreliable to be used as references. Even the raw read counts mappable to the reference spike in sequences were significantly out of the linear correlations between any two samples. As a workaround we picked the internal controls from the first set to find genes that express more than 500 normalized reads with ≤ 0.2 of the log2 largest difference between two samples. Fourteen genes satisfied the criteria to become a set of internal controls. The internal reference genes were used in place of spike in RNAs for processing counts by the modified indirect RUVg approach. Genome build: GRCz10 Supplementary files format and content: comma separated text file include RUVg normalized rs1a and rs1b or TPM normalized rs2 values for each sample. Zebrafish whole embryo Approximately 1 nl of the solution containing TUT4/7 or control morpholinos was injected into wild type zebrafish embryos at xxx cell stage. For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. Zebrafish embryos were obtained by the natural mating of wild type AB strain and grown in embryo medium at 28.5˚C. Embryos were staged according to standard morphological criteria. hpf GSM3024080 GSM3024080: Zebrafish 3 hpf whole embryo injected with ConMO Exp. ID: rs2; Danio rerio; RNA Seq GSM3024080   1 For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. GEO Accession:GSM3024080 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP133517     rs2-ConMO-3hpf_R1.fastq.gz rs2-ConMO-3hpf_R2.fastq.gz fastq fastq 7456939576.0 49058813.0 GSM3024080 r1 0:76 1:76 A:1813253495;C:1892010922;G:1885775058;T:1863713907;N:2186194 76 76     1813253495 1892010922 1885775058 1863713907 2186194 SRX3744226 SRS3000694 SRA662359 GEO Narry Kim Lab, Center for RNA Research, Institute for Basic Science 2 0.94906 0.95288 0.06949 0.06558 0.75749 0.75893 0.57185 0.59121 76 76 B B biological fallback assumption illumina nextseq unknown poly_a ribozero bulk bulk bulk   South Korea 2018-02-26 Blastula Embryo Whole Organism All anatomical structures
47622 47622 SRR6784686 SRX3744224 SRS3000698 SRP133517 PRJNA436017 Maternal mRNA degradation by TUT4 and TUT7 in the zebrafish maternal to zygotic transition GSE111152 Transcriptome Analysis During the maternal to zygotic transition MZT maternal RNAs are actively degraded and replaced by newly synthesized zygotic transcripts in a highly coordinated manner. However it remains largely unknown how maternal mRNA decay is triggered in early vertebrate embryos. Here through genome wide profiling of RNA abundance and three prime modification we show that uridylation is induced at the onset of maternal mRNA clearance. The temporal control of uridylation is conserved in vertebrates. When the homologs of terminal uridylyltransferases TUT4 and TUT7 TUT4/7 are depleted in zebrafish and Xenopus maternal mRNA clearance is significantly delayed leading to developmental defects during gastrulation. Short tailed mRNAs are selectively uridylated by TUT4/7 with the highly uridylated transcripts degraded faster during the MZT than those with unmodified polyA tails. Our study demonstrates that uridylation plays a crucial role in timely mRNA degradation thereby allowing the progression of early development. Overall design: Examination of zebrafish embryos injected with control or TUT4/7 morpholinos at 1 cell stage. RNA seq libraries were prepared at different time points across the MZT period. Three experimental sets rs1a rs2b and rs2 were conducted independently.   pubmed:29625039   Zebrafish 3 hpf whole embryo injected with TUT47MO Exp. ID: rs1b GSM3024078   tissue:Zebrafish whole embryo|hpf translation blocking TUT7 translation blocking|strain:AB Zebrafish 3 hpf whole embryo injected with TUT47MO Exp. ID: rs1b The sequences were basecalled by Illumina RTA embedded in NextSeq Control Software 1.3.2 by the standard workflow. Sequenced reads from RNA seq data were mapped to the GRCz10 genome with RefSeq release 104 annotations above using STAR 2.5.3a via RSEM 2.1.31. The gene level quantifications were performed using RSEM with the default options for strand specific and paired end RNA seq. The "rs1a" set of our libraries was between sample normalized using RUV seq 1.10.0. The standard workflow suggested by RUV seq did not work due to the global changes of maternal transcripts following the progression of embryogenesis. Instead we first chose a subset of spike in RNAs that have ≥ 256 “expected counts” of reads in at least 90% of samples. Then the counts were normalized by the RUVg algorithm k = 1. The geometric means of the normalized counts were used as size factors for the scale normalization of all other RNAs. The ERCC spike in reads in our second RNA seq dataset were too unreliable to be used as references. Even the raw read counts mappable to the reference spike in sequences were significantly out of the linear correlations between any two samples. As a workaround we picked the internal controls from the first set to find genes that express more than 500 normalized reads with ≤ 0.2 of the log2 largest difference between two samples. Fourteen genes satisfied the criteria to become a set of internal controls. The internal reference genes were used in place of spike in RNAs for processing counts by the modified indirect RUVg approach. Genome build: GRCz10 Supplementary files format and content: comma separated text file include RUVg normalized rs1a and rs1b or TPM normalized rs2 values for each sample. Zebrafish whole embryo Approximately 1 nl of the solution containing TUT4/7 or control morpholinos was injected into wild type zebrafish embryos at xxx cell stage. For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. Zebrafish embryos were obtained by the natural mating of wild type AB strain and grown in embryo medium at 28.5˚C. Embryos were staged according to standard morphological criteria. hpf translation blocking TUT7 translation blocking|strain:AB GSM3024078 GSM3024078: Zebrafish 3 hpf whole embryo injected with TUT47MO Exp. ID: rs1b; Danio rerio; RNA Seq GSM3024078   1 For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. GEO Accession:GSM3024078 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP133517     rs1b-TUT47MO-3hpf_R1.fastq.gz rs1b-TUT47MO-3hpf_R2.fastq.gz fastq fastq 3512955750.0 23419705.0 GSM3024078 r1 0:75 1:75 A:904272993;C:846189136;G:845724027;T:914848835;N:1920759 75 75     904272993 846189136 845724027 914848835 1920759 SRX3744224 SRS3000698 SRA662359 GEO Narry Kim Lab, Center for RNA Research, Institute for Basic Science 2 0.94996 0.95308 0.06884 0.06709 0.74546 0.74856 0.49941 0.49729 75 75 B B biological fallback assumption illumina nextseq unknown poly_a ribozero bulk bulk bulk   South Korea 2018-02-26 Blastula Embryo Whole Organism All anatomical structures
47624 47624 SRR6784684 SRX3744222 SRS3000691 SRP133517 PRJNA436017 Maternal mRNA degradation by TUT4 and TUT7 in the zebrafish maternal to zygotic transition GSE111152 Transcriptome Analysis During the maternal to zygotic transition MZT maternal RNAs are actively degraded and replaced by newly synthesized zygotic transcripts in a highly coordinated manner. However it remains largely unknown how maternal mRNA decay is triggered in early vertebrate embryos. Here through genome wide profiling of RNA abundance and three prime modification we show that uridylation is induced at the onset of maternal mRNA clearance. The temporal control of uridylation is conserved in vertebrates. When the homologs of terminal uridylyltransferases TUT4 and TUT7 TUT4/7 are depleted in zebrafish and Xenopus maternal mRNA clearance is significantly delayed leading to developmental defects during gastrulation. Short tailed mRNAs are selectively uridylated by TUT4/7 with the highly uridylated transcripts degraded faster during the MZT than those with unmodified polyA tails. Our study demonstrates that uridylation plays a crucial role in timely mRNA degradation thereby allowing the progression of early development. Overall design: Examination of zebrafish embryos injected with control or TUT4/7 morpholinos at 1 cell stage. RNA seq libraries were prepared at different time points across the MZT period. Three experimental sets rs1a rs2b and rs2 were conducted independently.   pubmed:29625039   Zebrafish 3 hpf whole embryo injected with ConMO Exp. ID: rs1b GSM3024076   tissue:Zebrafish whole embryo|hpf Zebrafish 3 hpf whole embryo injected with ConMO Exp. ID: rs1b The sequences were basecalled by Illumina RTA embedded in NextSeq Control Software 1.3.2 by the standard workflow. Sequenced reads from RNA seq data were mapped to the GRCz10 genome with RefSeq release 104 annotations above using STAR 2.5.3a via RSEM 2.1.31. The gene level quantifications were performed using RSEM with the default options for strand specific and paired end RNA seq. The "rs1a" set of our libraries was between sample normalized using RUV seq 1.10.0. The standard workflow suggested by RUV seq did not work due to the global changes of maternal transcripts following the progression of embryogenesis. Instead we first chose a subset of spike in RNAs that have ≥ 256 “expected counts” of reads in at least 90% of samples. Then the counts were normalized by the RUVg algorithm k = 1. The geometric means of the normalized counts were used as size factors for the scale normalization of all other RNAs. The ERCC spike in reads in our second RNA seq dataset were too unreliable to be used as references. Even the raw read counts mappable to the reference spike in sequences were significantly out of the linear correlations between any two samples. As a workaround we picked the internal controls from the first set to find genes that express more than 500 normalized reads with ≤ 0.2 of the log2 largest difference between two samples. Fourteen genes satisfied the criteria to become a set of internal controls. The internal reference genes were used in place of spike in RNAs for processing counts by the modified indirect RUVg approach. Genome build: GRCz10 Supplementary files format and content: comma separated text file include RUVg normalized rs1a and rs1b or TPM normalized rs2 values for each sample. Zebrafish whole embryo Approximately 1 nl of the solution containing TUT4/7 or control morpholinos was injected into wild type zebrafish embryos at xxx cell stage. For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. Zebrafish embryos were obtained by the natural mating of wild type AB strain and grown in embryo medium at 28.5˚C. Embryos were staged according to standard morphological criteria. hpf GSM3024076 GSM3024076: Zebrafish 3 hpf whole embryo injected with ConMO Exp. ID: rs1b; Danio rerio; RNA Seq GSM3024076   1 For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. GEO Accession:GSM3024076 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP133517     rs1b-ConMO-3hpf_R1.fastq.gz rs1b-ConMO-3hpf_R2.fastq.gz fastq fastq 3588228600.0 23921524.0 GSM3024076 r1 0:75 1:75 A:927704680;C:861896753;G:859653360;T:937028832;N:1944975 75 75     927704680 861896753 859653360 937028832 1944975 SRX3744222 SRS3000691 SRA662359 GEO Narry Kim Lab, Center for RNA Research, Institute for Basic Science 2 0.9482 0.95265 0.06761 0.06634 0.74673 0.74949 0.5033 0.50552 75 75 B B biological fallback assumption illumina nextseq unknown poly_a ribozero bulk bulk bulk   South Korea 2018-02-26 Blastula Embryo Whole Organism All anatomical structures
47626 47626 SRR6784682 SRX3744220 SRS3000689 SRP133517 PRJNA436017 Maternal mRNA degradation by TUT4 and TUT7 in the zebrafish maternal to zygotic transition GSE111152 Transcriptome Analysis During the maternal to zygotic transition MZT maternal RNAs are actively degraded and replaced by newly synthesized zygotic transcripts in a highly coordinated manner. However it remains largely unknown how maternal mRNA decay is triggered in early vertebrate embryos. Here through genome wide profiling of RNA abundance and three prime modification we show that uridylation is induced at the onset of maternal mRNA clearance. The temporal control of uridylation is conserved in vertebrates. When the homologs of terminal uridylyltransferases TUT4 and TUT7 TUT4/7 are depleted in zebrafish and Xenopus maternal mRNA clearance is significantly delayed leading to developmental defects during gastrulation. Short tailed mRNAs are selectively uridylated by TUT4/7 with the highly uridylated transcripts degraded faster during the MZT than those with unmodified polyA tails. Our study demonstrates that uridylation plays a crucial role in timely mRNA degradation thereby allowing the progression of early development. Overall design: Examination of zebrafish embryos injected with control or TUT4/7 morpholinos at 1 cell stage. RNA seq libraries were prepared at different time points across the MZT period. Three experimental sets rs1a rs2b and rs2 were conducted independently.   pubmed:29625039   Zebrafish 4 hpf whole embryo injected with TUT47MO Exp. ID: rs1a GSM3024074   tissue:Zebrafish whole embryo|hpf translation blocking TUT7 translation blocking|strain:AB Zebrafish 4 hpf whole embryo injected with TUT47MO Exp. ID: rs1a The sequences were basecalled by Illumina RTA embedded in NextSeq Control Software 1.3.2 by the standard workflow. Sequenced reads from RNA seq data were mapped to the GRCz10 genome with RefSeq release 104 annotations above using STAR 2.5.3a via RSEM 2.1.31. The gene level quantifications were performed using RSEM with the default options for strand specific and paired end RNA seq. The "rs1a" set of our libraries was between sample normalized using RUV seq 1.10.0. The standard workflow suggested by RUV seq did not work due to the global changes of maternal transcripts following the progression of embryogenesis. Instead we first chose a subset of spike in RNAs that have ≥ 256 “expected counts” of reads in at least 90% of samples. Then the counts were normalized by the RUVg algorithm k = 1. The geometric means of the normalized counts were used as size factors for the scale normalization of all other RNAs. The ERCC spike in reads in our second RNA seq dataset were too unreliable to be used as references. Even the raw read counts mappable to the reference spike in sequences were significantly out of the linear correlations between any two samples. As a workaround we picked the internal controls from the first set to find genes that express more than 500 normalized reads with ≤ 0.2 of the log2 largest difference between two samples. Fourteen genes satisfied the criteria to become a set of internal controls. The internal reference genes were used in place of spike in RNAs for processing counts by the modified indirect RUVg approach. Genome build: GRCz10 Supplementary files format and content: comma separated text file include RUVg normalized rs1a and rs1b or TPM normalized rs2 values for each sample. Zebrafish whole embryo Approximately 1 nl of the solution containing TUT4/7 or control morpholinos was injected into wild type zebrafish embryos at xxx cell stage. For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. Zebrafish embryos were obtained by the natural mating of wild type AB strain and grown in embryo medium at 28.5˚C. Embryos were staged according to standard morphological criteria. hpf translation blocking TUT7 translation blocking|strain:AB GSM3024074 GSM3024074: Zebrafish 4 hpf whole embryo injected with TUT47MO Exp. ID: rs1a; Danio rerio; RNA Seq GSM3024074   1 For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. GEO Accession:GSM3024074 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP133517     rs1a-TUT47MO-4hpf_R1.fastq.gz rs1a-TUT47MO-4hpf_R2.fastq.gz fastq fastq 4012253850.0 26748359.0 GSM3024074 r1 0:75 1:75 A:1083176195;C:921754621;G:910925804;T:1094191077;N:2206153 75 75     1083176195 921754621 910925804 1094191077 2206153 SRX3744220 SRS3000689 SRA662359 GEO Narry Kim Lab, Center for RNA Research, Institute for Basic Science 2 0.95565 0.96284 0.08391 0.08319 0.75923 0.76098 0.72617 0.72687 75 75 B B biological fallback assumption illumina nextseq unknown poly_a ribozero bulk bulk bulk   South Korea 2018-02-26 Blastula Embryo Whole Organism All anatomical structures
47629 47629 SRR6784679 SRX3744217 SRS3000686 SRP133517 PRJNA436017 Maternal mRNA degradation by TUT4 and TUT7 in the zebrafish maternal to zygotic transition GSE111152 Transcriptome Analysis During the maternal to zygotic transition MZT maternal RNAs are actively degraded and replaced by newly synthesized zygotic transcripts in a highly coordinated manner. However it remains largely unknown how maternal mRNA decay is triggered in early vertebrate embryos. Here through genome wide profiling of RNA abundance and three prime modification we show that uridylation is induced at the onset of maternal mRNA clearance. The temporal control of uridylation is conserved in vertebrates. When the homologs of terminal uridylyltransferases TUT4 and TUT7 TUT4/7 are depleted in zebrafish and Xenopus maternal mRNA clearance is significantly delayed leading to developmental defects during gastrulation. Short tailed mRNAs are selectively uridylated by TUT4/7 with the highly uridylated transcripts degraded faster during the MZT than those with unmodified polyA tails. Our study demonstrates that uridylation plays a crucial role in timely mRNA degradation thereby allowing the progression of early development. Overall design: Examination of zebrafish embryos injected with control or TUT4/7 morpholinos at 1 cell stage. RNA seq libraries were prepared at different time points across the MZT period. Three experimental sets rs1a rs2b and rs2 were conducted independently.   pubmed:29625039   Zebrafish 4 hpf whole embryo injected with TUT7MO Exp. ID: rs1a GSM3024071   tissue:Zebrafish whole embryo|hpf translation blocking|strain:AB Zebrafish 4 hpf whole embryo injected with TUT7MO Exp. ID: rs1a The sequences were basecalled by Illumina RTA embedded in NextSeq Control Software 1.3.2 by the standard workflow. Sequenced reads from RNA seq data were mapped to the GRCz10 genome with RefSeq release 104 annotations above using STAR 2.5.3a via RSEM 2.1.31. The gene level quantifications were performed using RSEM with the default options for strand specific and paired end RNA seq. The "rs1a" set of our libraries was between sample normalized using RUV seq 1.10.0. The standard workflow suggested by RUV seq did not work due to the global changes of maternal transcripts following the progression of embryogenesis. Instead we first chose a subset of spike in RNAs that have ≥ 256 “expected counts” of reads in at least 90% of samples. Then the counts were normalized by the RUVg algorithm k = 1. The geometric means of the normalized counts were used as size factors for the scale normalization of all other RNAs. The ERCC spike in reads in our second RNA seq dataset were too unreliable to be used as references. Even the raw read counts mappable to the reference spike in sequences were significantly out of the linear correlations between any two samples. As a workaround we picked the internal controls from the first set to find genes that express more than 500 normalized reads with ≤ 0.2 of the log2 largest difference between two samples. Fourteen genes satisfied the criteria to become a set of internal controls. The internal reference genes were used in place of spike in RNAs for processing counts by the modified indirect RUVg approach. Genome build: GRCz10 Supplementary files format and content: comma separated text file include RUVg normalized rs1a and rs1b or TPM normalized rs2 values for each sample. Zebrafish whole embryo Approximately 1 nl of the solution containing TUT4/7 or control morpholinos was injected into wild type zebrafish embryos at xxx cell stage. For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. Zebrafish embryos were obtained by the natural mating of wild type AB strain and grown in embryo medium at 28.5˚C. Embryos were staged according to standard morphological criteria. hpf translation blocking|strain:AB GSM3024071 GSM3024071: Zebrafish 4 hpf whole embryo injected with TUT7MO Exp. ID: rs1a; Danio rerio; RNA Seq GSM3024071   1 For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. GEO Accession:GSM3024071 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP133517     rs1a-TUT7MO-4hpf_R2.fastq.gz rs1a-TUT7MO-4hpf_R1.fastq.gz fastq fastq 5768921100.0 38459474.0 GSM3024071 r1 0:75 1:75 A:1555172603;C:1327529056;G:1315409284;T:1567647314;N:3162843 75 75     1555172603 1327529056 1315409284 1567647314 3162843 SRX3744217 SRS3000686 SRA662359 GEO Narry Kim Lab, Center for RNA Research, Institute for Basic Science 2 0.95735 0.96309 0.08315 0.08265 0.76493 0.76806 0.72767 0.73332 75 75 B B biological fallback assumption illumina nextseq unknown poly_a ribozero bulk bulk bulk   South Korea 2018-02-26 Blastula Embryo Whole Organism All anatomical structures
47632 47632 SRR6784676 SRX3744214 SRS3000683 SRP133517 PRJNA436017 Maternal mRNA degradation by TUT4 and TUT7 in the zebrafish maternal to zygotic transition GSE111152 Transcriptome Analysis During the maternal to zygotic transition MZT maternal RNAs are actively degraded and replaced by newly synthesized zygotic transcripts in a highly coordinated manner. However it remains largely unknown how maternal mRNA decay is triggered in early vertebrate embryos. Here through genome wide profiling of RNA abundance and three prime modification we show that uridylation is induced at the onset of maternal mRNA clearance. The temporal control of uridylation is conserved in vertebrates. When the homologs of terminal uridylyltransferases TUT4 and TUT7 TUT4/7 are depleted in zebrafish and Xenopus maternal mRNA clearance is significantly delayed leading to developmental defects during gastrulation. Short tailed mRNAs are selectively uridylated by TUT4/7 with the highly uridylated transcripts degraded faster during the MZT than those with unmodified polyA tails. Our study demonstrates that uridylation plays a crucial role in timely mRNA degradation thereby allowing the progression of early development. Overall design: Examination of zebrafish embryos injected with control or TUT4/7 morpholinos at 1 cell stage. RNA seq libraries were prepared at different time points across the MZT period. Three experimental sets rs1a rs2b and rs2 were conducted independently.   pubmed:29625039   Zebrafish 4 hpf whole embryo injected with TUT4MO Exp. ID: rs1a GSM3024068   tissue:Zebrafish whole embryo|hpf translation blocking|strain:AB Zebrafish 4 hpf whole embryo injected with TUT4MO Exp. ID: rs1a The sequences were basecalled by Illumina RTA embedded in NextSeq Control Software 1.3.2 by the standard workflow. Sequenced reads from RNA seq data were mapped to the GRCz10 genome with RefSeq release 104 annotations above using STAR 2.5.3a via RSEM 2.1.31. The gene level quantifications were performed using RSEM with the default options for strand specific and paired end RNA seq. The "rs1a" set of our libraries was between sample normalized using RUV seq 1.10.0. The standard workflow suggested by RUV seq did not work due to the global changes of maternal transcripts following the progression of embryogenesis. Instead we first chose a subset of spike in RNAs that have ≥ 256 “expected counts” of reads in at least 90% of samples. Then the counts were normalized by the RUVg algorithm k = 1. The geometric means of the normalized counts were used as size factors for the scale normalization of all other RNAs. The ERCC spike in reads in our second RNA seq dataset were too unreliable to be used as references. Even the raw read counts mappable to the reference spike in sequences were significantly out of the linear correlations between any two samples. As a workaround we picked the internal controls from the first set to find genes that express more than 500 normalized reads with ≤ 0.2 of the log2 largest difference between two samples. Fourteen genes satisfied the criteria to become a set of internal controls. The internal reference genes were used in place of spike in RNAs for processing counts by the modified indirect RUVg approach. Genome build: GRCz10 Supplementary files format and content: comma separated text file include RUVg normalized rs1a and rs1b or TPM normalized rs2 values for each sample. Zebrafish whole embryo Approximately 1 nl of the solution containing TUT4/7 or control morpholinos was injected into wild type zebrafish embryos at xxx cell stage. For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. Zebrafish embryos were obtained by the natural mating of wild type AB strain and grown in embryo medium at 28.5˚C. Embryos were staged according to standard morphological criteria. hpf translation blocking|strain:AB GSM3024068 GSM3024068: Zebrafish 4 hpf whole embryo injected with TUT4MO Exp. ID: rs1a; Danio rerio; RNA Seq GSM3024068   1 For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. GEO Accession:GSM3024068 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP133517     rs1a-TUT4MO-4hpf_R2.fastq.gz rs1a-TUT4MO-4hpf_R1.fastq.gz fastq fastq 4215564300.0 28103762.0 GSM3024068 r1 0:75 1:75 A:1065121902;C:1041520955;G:1038600008;T:1068046442;N:2274993 75 75     1065121902 1041520955 1038600008 1068046442 2274993 SRX3744214 SRS3000683 SRA662359 GEO Narry Kim Lab, Center for RNA Research, Institute for Basic Science 2 0.94653 0.95013 0.10969 0.10755 0.75469 0.75777 0.64019 0.6461 75 75 B B biological fallback assumption illumina nextseq unknown poly_a ribozero bulk bulk bulk   South Korea 2018-02-26 Blastula Embryo Whole Organism All anatomical structures
47635 47635 SRR6784673 SRX3744211 SRS3000680 SRP133517 PRJNA436017 Maternal mRNA degradation by TUT4 and TUT7 in the zebrafish maternal to zygotic transition GSE111152 Transcriptome Analysis During the maternal to zygotic transition MZT maternal RNAs are actively degraded and replaced by newly synthesized zygotic transcripts in a highly coordinated manner. However it remains largely unknown how maternal mRNA decay is triggered in early vertebrate embryos. Here through genome wide profiling of RNA abundance and three prime modification we show that uridylation is induced at the onset of maternal mRNA clearance. The temporal control of uridylation is conserved in vertebrates. When the homologs of terminal uridylyltransferases TUT4 and TUT7 TUT4/7 are depleted in zebrafish and Xenopus maternal mRNA clearance is significantly delayed leading to developmental defects during gastrulation. Short tailed mRNAs are selectively uridylated by TUT4/7 with the highly uridylated transcripts degraded faster during the MZT than those with unmodified polyA tails. Our study demonstrates that uridylation plays a crucial role in timely mRNA degradation thereby allowing the progression of early development. Overall design: Examination of zebrafish embryos injected with control or TUT4/7 morpholinos at 1 cell stage. RNA seq libraries were prepared at different time points across the MZT period. Three experimental sets rs1a rs2b and rs2 were conducted independently.   pubmed:29625039   Zebrafish 4 hpf whole embryo injected with ConMO Exp. ID: rs1a GSM3024065   tissue:Zebrafish whole embryo|hpf Zebrafish 4 hpf whole embryo injected with ConMO Exp. ID: rs1a The sequences were basecalled by Illumina RTA embedded in NextSeq Control Software 1.3.2 by the standard workflow. Sequenced reads from RNA seq data were mapped to the GRCz10 genome with RefSeq release 104 annotations above using STAR 2.5.3a via RSEM 2.1.31. The gene level quantifications were performed using RSEM with the default options for strand specific and paired end RNA seq. The "rs1a" set of our libraries was between sample normalized using RUV seq 1.10.0. The standard workflow suggested by RUV seq did not work due to the global changes of maternal transcripts following the progression of embryogenesis. Instead we first chose a subset of spike in RNAs that have ≥ 256 “expected counts” of reads in at least 90% of samples. Then the counts were normalized by the RUVg algorithm k = 1. The geometric means of the normalized counts were used as size factors for the scale normalization of all other RNAs. The ERCC spike in reads in our second RNA seq dataset were too unreliable to be used as references. Even the raw read counts mappable to the reference spike in sequences were significantly out of the linear correlations between any two samples. As a workaround we picked the internal controls from the first set to find genes that express more than 500 normalized reads with ≤ 0.2 of the log2 largest difference between two samples. Fourteen genes satisfied the criteria to become a set of internal controls. The internal reference genes were used in place of spike in RNAs for processing counts by the modified indirect RUVg approach. Genome build: GRCz10 Supplementary files format and content: comma separated text file include RUVg normalized rs1a and rs1b or TPM normalized rs2 values for each sample. Zebrafish whole embryo Approximately 1 nl of the solution containing TUT4/7 or control morpholinos was injected into wild type zebrafish embryos at xxx cell stage. For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. Zebrafish embryos were obtained by the natural mating of wild type AB strain and grown in embryo medium at 28.5˚C. Embryos were staged according to standard morphological criteria. hpf GSM3024065 GSM3024065: Zebrafish 4 hpf whole embryo injected with ConMO Exp. ID: rs1a; Danio rerio; RNA Seq GSM3024065   1 For each sample embryos were collected and total RNA was extracted with TRIzol Invitrogen. The samples were spiked in with ERCC RNA Spike In Mix Ambion for normalization purpose. The quality of total RNA was checked by Agilent Bioanalyzer 2100 Agilent. Ribosomal RNA was depleted from total RNA using Ribo Zero Gold kit Epicentre. RNA seq libraries were constructed using TruSeq stranded total RNA library prep kit Illumina and sequenced in Illumina Nextseq 500 v2 to yield paired end 76nt reads. GEO Accession:GSM3024065 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP133517     rs1a-ConMO-4hpf_R2.fastq.gz rs1a-ConMO-4hpf_R1.fastq.gz fastq fastq 3861279450.0 25741863.0 GSM3024065 r1 0:75 1:75 A:973485072;C:956743877;G:950463164;T:978475358;N:2111979 75 75     973485072 956743877 950463164 978475358 2111979 SRX3744211 SRS3000680 SRA662359 GEO Narry Kim Lab, Center for RNA Research, Institute for Basic Science 2 0.9482 0.95259 0.11181 0.10881 0.7555 0.75763 0.64284 0.64083 75 75 B B biological fallback assumption illumina nextseq unknown poly_a ribozero bulk bulk bulk   South Korea 2018-02-26 Blastula Embryo Whole Organism All anatomical structures
53282 53282 SRR11869572 SRX8419562 SRS6731106 SRP216221 PRJNA556409 RNA seq of mondoa zebrafish mutants GSE134778 Transcriptome Analysis The glucose sensing Mondo pathway regulates expression of metabolic genes in mammals. Here we have revealed an unexpected role of this pathway in vertebrate embryonic development. We characterized Mondo pathway function in the zebrafish and showed that knock down of mondoa impaired the early morphogenetic movement of epib in zebrafish embryos. Expression of nsdhl an enzyme of cholesterol and pregnenolone synthesis was strongly reduced in these embryos. Loss of Nsdhl function likewise impaired epib and led to microtubule cytoskeleton defects in the embryo similar to MondoA loss of function. both epib and microtubule cytoskeleton defects were partially restored by pregnenolone treatment. Gene disruption of mondoa perturbed epib with only partial penetrance likely due to compensatory changes in the expression of cholesterol/steroid metabolism genes. Collectively our results show a novel role for MondoA in the regulation of early vertebrate development connecting glucose cholesterol and steroid hormone metabolism with early embryonic cell movements. Overall design: RNA seq in mondoa zebrafish mutants parent bioproject:PRJNA603464 pubmed:32969791   mondoa s 3 GSM4577399   source name:total RNA|tissue:embryo|genotype:MZmondoA mutant allel: ka405 ZFIN ID: ZDB ALT 180628 2|phenotype:affected epib|Stage:5 hpf mondoa s 3 Image analysis and base calling were performed using RTA 2.7.3 and bcl2fastq 2.17.1.14. Sequences were aligned to the full reference genome index build with Ensembl annotation to improve accuracy on splices junctions using STAR 2.4.0i A custom Perl script was used to quantify uniquely mapped reads. For each Ensembl gene all exons of the respective annotated protein coding transcripts were considered. Genome build: zv10 Supplementary files format and content: counts exon.txt exonic counts for each indicated gene and sample total RNA   Embryos were sampled in liquid nitrogen and stored at 80°C until further processing. RNA was extracted with TRIzol Life Technologies following the manufacturer’s instructions with minor modifications: 30 larvae were sampled in a volume of 1.5 ml TRIzol reagent. Samples were kept at least overnight at 80°C. In order to disrupt the cell membranes the samples were passed four times through a syringe with a diameter of 0.45 mm. Total RNA Seq libraries were generated from 500 ng of total RNA using TruSeq Stranded Total RNA Library Prep Human/Mouse/Rat kit and TruSeq RNA Single Indexes kits A and B Illumina San Diego CA according to manufacturer's instructions. Briefly cytoplasmic and mitochondrial ribosomal RNA rRNA was removed using biotinylated target specific oligos combined with Ribo Zero rRNA removal beads. Following purification the depleted RNA was fragmented into small pieces using divalent cations at 94oC for 2 minutes. Cleaved RNA fragments were then copied into first strand cDNA using reverse transcriptase and random primers followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. Strand specificity was achieved by replacing dTTP with dUTP during second strand synthesis. The double stranded cDNA fragments were blunted using T4 DNA polymerase Klenow DNA polymerase and T4 PNK. A single 'A' nucleotide was added to the three prime ends of the blunt DNA fragments using a Klenow fragment three prime to five primeexo minus enzyme. The cDNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR amplification 30 sec at 98oC; [10 sec at 98oC 30 sec at 60oC 30 sec at 72oC] x 12 cycles; 5 min at 72oC. Surplus PCR primers were further removed by purification using AMPure XP beads Beckman Coulter Villepinte France and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis.   tissue:embryo|genotype:MZmondoA mutant allel: ka405 ZFIN ID: ZDB ALT 180628 2|phenotype:affected epib|Stage:5 hpf GSM4577399 GSM4577399: F mondoa s 3; Danio rerio; RNA Seq GSM4577399   1 Embryos were sampled in liquid nitrogen and stored at 80°C until further processing. RNA was extracted with TRIzol Life Technologies following the manufacturer's instructions with minor modifications: 30 larvae were sampled in a volume of 1.5 ml TRIzol reagent. Samples were kept at least overnight at 80°C. In order to disrupt the cell membranes the samples were passed four times through a syringe with a diameter of 0.45 mm. Total RNA Seq libraries were generated from 500 ng of total RNA using TruSeq Stranded Total RNA Library Prep Human/Mouse/Rat kit and TruSeq RNA Single Indexes kits A and B Illumina San Diego CA according to manufacturer's instructions. Briefly cytoplasmic and mitochondrial ribosomal RNA rRNA was removed using biotinylated target specific oligos combined with Ribo Zero rRNA removal beads. Following purification the depleted RNA was fragmented into small pieces using divalent cations at 94oC for 2 minutes. Cleaved RNA fragments were then copied into first strand cDNA using reverse transcriptase and random primers followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. Strand specificity was achieved by replacing dTTP with dUTP during second strand synthesis. The double stranded cDNA fragments were blunted using T4 DNA polymerase Klenow DNA polymerase and T4 PNK. A single 'A' nucleotide was added to the three prime ends of the blunt DNA fragments using a Klenow fragment three prime to five primeexo minus enzyme. The cDNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR amplification 30 sec at 98oC; [10 sec at 98oC 30 sec at 60oC 30 sec at 72oC] x 12 cycles; 5 min at 72oC. Surplus PCR primers were further removed by purification using AMPure XP beads Beckman Coulter Villepinte France and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis. GEO Accession:GSM4577399 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP216221     VRGN25_S2.R1.fastq VRGN25_S2.R2.fastq fastq fastq 7551918900.0 75519189.0 GSM4577399 r1 0:50 1:50 A:2066041718;C:1702385113;G:1686618839;T:2095697396;N:1175834 50 50     2066041718 1702385113 1686618839 2095697396 1175834 SRX8419562 SRS6731106 SRA925866 GEO Naef Lab, SV, EPFL 2 0.8972 0.89917 0.27419 0.27338 0.74576 0.75668 0.50768 0.5128 50 50 B B biological fallback assumption illumina hiseq_era 3prime rrna_depletion trueseq bulk bulk bulk   Switzerland 2020-05-28 Blastula Embryo Embryo Imprecise All anatomical structures
53283 53283 SRR11869571 SRX8419561 SRS6731105 SRP216221 PRJNA556409 RNA seq of mondoa zebrafish mutants GSE134778 Transcriptome Analysis The glucose sensing Mondo pathway regulates expression of metabolic genes in mammals. Here we have revealed an unexpected role of this pathway in vertebrate embryonic development. We characterized Mondo pathway function in the zebrafish and showed that knock down of mondoa impaired the early morphogenetic movement of epib in zebrafish embryos. Expression of nsdhl an enzyme of cholesterol and pregnenolone synthesis was strongly reduced in these embryos. Loss of Nsdhl function likewise impaired epib and led to microtubule cytoskeleton defects in the embryo similar to MondoA loss of function. both epib and microtubule cytoskeleton defects were partially restored by pregnenolone treatment. Gene disruption of mondoa perturbed epib with only partial penetrance likely due to compensatory changes in the expression of cholesterol/steroid metabolism genes. Collectively our results show a novel role for MondoA in the regulation of early vertebrate development connecting glucose cholesterol and steroid hormone metabolism with early embryonic cell movements. Overall design: RNA seq in mondoa zebrafish mutants parent bioproject:PRJNA603464 pubmed:32969791   mondoa s 2 GSM4577398   source name:total RNA|tissue:embryo|genotype:MZmondoA mutant allel: ka405 ZFIN ID: ZDB ALT 180628 2|phenotype:affected epib|Stage:5 hpf mondoa s 2 Image analysis and base calling were performed using RTA 2.7.3 and bcl2fastq 2.17.1.14. Sequences were aligned to the full reference genome index build with Ensembl annotation to improve accuracy on splices junctions using STAR 2.4.0i A custom Perl script was used to quantify uniquely mapped reads. For each Ensembl gene all exons of the respective annotated protein coding transcripts were considered. Genome build: zv10 Supplementary files format and content: counts exon.txt exonic counts for each indicated gene and sample total RNA   Embryos were sampled in liquid nitrogen and stored at 80°C until further processing. RNA was extracted with TRIzol Life Technologies following the manufacturer’s instructions with minor modifications: 30 larvae were sampled in a volume of 1.5 ml TRIzol reagent. Samples were kept at least overnight at 80°C. In order to disrupt the cell membranes the samples were passed four times through a syringe with a diameter of 0.45 mm. Total RNA Seq libraries were generated from 500 ng of total RNA using TruSeq Stranded Total RNA Library Prep Human/Mouse/Rat kit and TruSeq RNA Single Indexes kits A and B Illumina San Diego CA according to manufacturer's instructions. Briefly cytoplasmic and mitochondrial ribosomal RNA rRNA was removed using biotinylated target specific oligos combined with Ribo Zero rRNA removal beads. Following purification the depleted RNA was fragmented into small pieces using divalent cations at 94oC for 2 minutes. Cleaved RNA fragments were then copied into first strand cDNA using reverse transcriptase and random primers followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. Strand specificity was achieved by replacing dTTP with dUTP during second strand synthesis. The double stranded cDNA fragments were blunted using T4 DNA polymerase Klenow DNA polymerase and T4 PNK. A single 'A' nucleotide was added to the three prime ends of the blunt DNA fragments using a Klenow fragment three prime to five primeexo minus enzyme. The cDNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR amplification 30 sec at 98oC; [10 sec at 98oC 30 sec at 60oC 30 sec at 72oC] x 12 cycles; 5 min at 72oC. Surplus PCR primers were further removed by purification using AMPure XP beads Beckman Coulter Villepinte France and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis.   tissue:embryo|genotype:MZmondoA mutant allel: ka405 ZFIN ID: ZDB ALT 180628 2|phenotype:affected epib|Stage:5 hpf GSM4577398 GSM4577398: E mondoa s 2; Danio rerio; RNA Seq GSM4577398   1 Embryos were sampled in liquid nitrogen and stored at 80°C until further processing. RNA was extracted with TRIzol Life Technologies following the manufacturer's instructions with minor modifications: 30 larvae were sampled in a volume of 1.5 ml TRIzol reagent. Samples were kept at least overnight at 80°C. In order to disrupt the cell membranes the samples were passed four times through a syringe with a diameter of 0.45 mm. Total RNA Seq libraries were generated from 500 ng of total RNA using TruSeq Stranded Total RNA Library Prep Human/Mouse/Rat kit and TruSeq RNA Single Indexes kits A and B Illumina San Diego CA according to manufacturer's instructions. Briefly cytoplasmic and mitochondrial ribosomal RNA rRNA was removed using biotinylated target specific oligos combined with Ribo Zero rRNA removal beads. Following purification the depleted RNA was fragmented into small pieces using divalent cations at 94oC for 2 minutes. Cleaved RNA fragments were then copied into first strand cDNA using reverse transcriptase and random primers followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. Strand specificity was achieved by replacing dTTP with dUTP during second strand synthesis. The double stranded cDNA fragments were blunted using T4 DNA polymerase Klenow DNA polymerase and T4 PNK. A single 'A' nucleotide was added to the three prime ends of the blunt DNA fragments using a Klenow fragment three prime to five primeexo minus enzyme. The cDNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR amplification 30 sec at 98oC; [10 sec at 98oC 30 sec at 60oC 30 sec at 72oC] x 12 cycles; 5 min at 72oC. Surplus PCR primers were further removed by purification using AMPure XP beads Beckman Coulter Villepinte France and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis. GEO Accession:GSM4577398 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP216221     VRGN24_S2.R1.fastq VRGN24_S2.R2.fastq fastq fastq 7666458300.0 76664583.0 GSM4577398 r1 0:50 1:50 A:2108884078;C:1705527841;G:1677597808;T:2173267709;N:1180864 50 50     2108884078 1705527841 1677597808 2173267709 1180864 SRX8419561 SRS6731105 SRA925866 GEO Naef Lab, SV, EPFL 2 0.89319 0.89501 0.29973 0.29906 0.74931 0.75724 0.50887 0.50894 50 50 B B biological fallback assumption illumina hiseq_era 3prime rrna_depletion trueseq bulk bulk bulk   Switzerland 2020-05-28 Blastula Embryo Embryo Imprecise All anatomical structures
53284 53284 SRR11869570 SRX8419560 SRS6731104 SRP216221 PRJNA556409 RNA seq of mondoa zebrafish mutants GSE134778 Transcriptome Analysis The glucose sensing Mondo pathway regulates expression of metabolic genes in mammals. Here we have revealed an unexpected role of this pathway in vertebrate embryonic development. We characterized Mondo pathway function in the zebrafish and showed that knock down of mondoa impaired the early morphogenetic movement of epib in zebrafish embryos. Expression of nsdhl an enzyme of cholesterol and pregnenolone synthesis was strongly reduced in these embryos. Loss of Nsdhl function likewise impaired epib and led to microtubule cytoskeleton defects in the embryo similar to MondoA loss of function. both epib and microtubule cytoskeleton defects were partially restored by pregnenolone treatment. Gene disruption of mondoa perturbed epib with only partial penetrance likely due to compensatory changes in the expression of cholesterol/steroid metabolism genes. Collectively our results show a novel role for MondoA in the regulation of early vertebrate development connecting glucose cholesterol and steroid hormone metabolism with early embryonic cell movements. Overall design: RNA seq in mondoa zebrafish mutants parent bioproject:PRJNA603464 pubmed:32969791   mondoa s 1 GSM4577397   source name:total RNA|tissue:embryo|genotype:MZmondoA mutant allel: ka405 ZFIN ID: ZDB ALT 180628 2|phenotype:affected epib|Stage:5 hpf mondoa s 1 Image analysis and base calling were performed using RTA 2.7.3 and bcl2fastq 2.17.1.14. Sequences were aligned to the full reference genome index build with Ensembl annotation to improve accuracy on splices junctions using STAR 2.4.0i A custom Perl script was used to quantify uniquely mapped reads. For each Ensembl gene all exons of the respective annotated protein coding transcripts were considered. Genome build: zv10 Supplementary files format and content: counts exon.txt exonic counts for each indicated gene and sample total RNA   Embryos were sampled in liquid nitrogen and stored at 80°C until further processing. RNA was extracted with TRIzol Life Technologies following the manufacturer’s instructions with minor modifications: 30 larvae were sampled in a volume of 1.5 ml TRIzol reagent. Samples were kept at least overnight at 80°C. In order to disrupt the cell membranes the samples were passed four times through a syringe with a diameter of 0.45 mm. Total RNA Seq libraries were generated from 500 ng of total RNA using TruSeq Stranded Total RNA Library Prep Human/Mouse/Rat kit and TruSeq RNA Single Indexes kits A and B Illumina San Diego CA according to manufacturer's instructions. Briefly cytoplasmic and mitochondrial ribosomal RNA rRNA was removed using biotinylated target specific oligos combined with Ribo Zero rRNA removal beads. Following purification the depleted RNA was fragmented into small pieces using divalent cations at 94oC for 2 minutes. Cleaved RNA fragments were then copied into first strand cDNA using reverse transcriptase and random primers followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. Strand specificity was achieved by replacing dTTP with dUTP during second strand synthesis. The double stranded cDNA fragments were blunted using T4 DNA polymerase Klenow DNA polymerase and T4 PNK. A single 'A' nucleotide was added to the three prime ends of the blunt DNA fragments using a Klenow fragment three prime to five primeexo minus enzyme. The cDNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR amplification 30 sec at 98oC; [10 sec at 98oC 30 sec at 60oC 30 sec at 72oC] x 12 cycles; 5 min at 72oC. Surplus PCR primers were further removed by purification using AMPure XP beads Beckman Coulter Villepinte France and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis.   tissue:embryo|genotype:MZmondoA mutant allel: ka405 ZFIN ID: ZDB ALT 180628 2|phenotype:affected epib|Stage:5 hpf GSM4577397 GSM4577397: D mondoa s 1; Danio rerio; RNA Seq GSM4577397   1 Embryos were sampled in liquid nitrogen and stored at 80°C until further processing. RNA was extracted with TRIzol Life Technologies following the manufacturer's instructions with minor modifications: 30 larvae were sampled in a volume of 1.5 ml TRIzol reagent. Samples were kept at least overnight at 80°C. In order to disrupt the cell membranes the samples were passed four times through a syringe with a diameter of 0.45 mm. Total RNA Seq libraries were generated from 500 ng of total RNA using TruSeq Stranded Total RNA Library Prep Human/Mouse/Rat kit and TruSeq RNA Single Indexes kits A and B Illumina San Diego CA according to manufacturer's instructions. Briefly cytoplasmic and mitochondrial ribosomal RNA rRNA was removed using biotinylated target specific oligos combined with Ribo Zero rRNA removal beads. Following purification the depleted RNA was fragmented into small pieces using divalent cations at 94oC for 2 minutes. Cleaved RNA fragments were then copied into first strand cDNA using reverse transcriptase and random primers followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. Strand specificity was achieved by replacing dTTP with dUTP during second strand synthesis. The double stranded cDNA fragments were blunted using T4 DNA polymerase Klenow DNA polymerase and T4 PNK. A single 'A' nucleotide was added to the three prime ends of the blunt DNA fragments using a Klenow fragment three prime to five primeexo minus enzyme. The cDNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR amplification 30 sec at 98oC; [10 sec at 98oC 30 sec at 60oC 30 sec at 72oC] x 12 cycles; 5 min at 72oC. Surplus PCR primers were further removed by purification using AMPure XP beads Beckman Coulter Villepinte France and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis. GEO Accession:GSM4577397 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP216221     VRGN23_S2.R1.fastq VRGN23_S2.R2.fastq fastq fastq 6872302500.0 68723025.0 GSM4577397 r1 0:50 1:50 A:1873323778;C:1557060731;G:1546029001;T:1894832528;N:1056462 50 50     1873323778 1557060731 1546029001 1894832528 1056462 SRX8419560 SRS6731104 SRA925866 GEO Naef Lab, SV, EPFL 2 0.89976 0.90345 0.27147 0.27139 0.74957 0.76187 0.50583 0.51168 50 50 B B biological fallback assumption illumina hiseq_era 3prime rrna_depletion trueseq bulk bulk bulk   Switzerland 2020-05-28 Blastula Embryo Embryo Imprecise All anatomical structures
53285 53285 SRR9822880 SRX6579572 SRS5145283 SRP216221 PRJNA556409 RNA seq of mondoa zebrafish mutants GSE134778 Transcriptome Analysis The glucose sensing Mondo pathway regulates expression of metabolic genes in mammals. Here we have revealed an unexpected role of this pathway in vertebrate embryonic development. We characterized Mondo pathway function in the zebrafish and showed that knock down of mondoa impaired the early morphogenetic movement of epib in zebrafish embryos. Expression of nsdhl an enzyme of cholesterol and pregnenolone synthesis was strongly reduced in these embryos. Loss of Nsdhl function likewise impaired epib and led to microtubule cytoskeleton defects in the embryo similar to MondoA loss of function. both epib and microtubule cytoskeleton defects were partially restored by pregnenolone treatment. Gene disruption of mondoa perturbed epib with only partial penetrance likely due to compensatory changes in the expression of cholesterol/steroid metabolism genes. Collectively our results show a novel role for MondoA in the regulation of early vertebrate development connecting glucose cholesterol and steroid hormone metabolism with early embryonic cell movements. Overall design: RNA seq in mondoa zebrafish mutants parent bioproject:PRJNA603464 pubmed:32969791   WT n 3 GSM3968576   source name:total RNA|tissue:embryo|genotype:wild type|phenotype:unaffected epib|Stage:5 hpf WT n 3 Image analysis and base calling were performed using RTA 2.7.3 and bcl2fastq 2.17.1.14. Sequences were aligned to the full reference genome index build with Ensembl annotation to improve accuracy on splices junctions using STAR 2.4.0i A custom Perl script was used to quantify uniquely mapped reads. For each Ensembl gene all exons of the respective annotated protein coding transcripts were considered. Genome build: zv10 Supplementary files format and content: counts mutants.txt: Tab delimited text file contains raw counts. total RNA   Embryos were sampled in liquid nitrogen and stored at 80°C until further processing. RNA was extracted with TRIzol Life Technologies following the manufacturer’s instructions with minor modifications: 30 larvae were sampled in a volume of 1.5 ml TRIzol reagent. Samples were kept at least overnight at 80°C. In order to disrupt the cell membranes the samples were passed four times through a syringe with a diameter of 0.45 mm. Total RNA Seq libraries were generated from 500 ng of total RNA using TruSeq Stranded Total RNA Library Prep Human/Mouse/Rat kit and TruSeq RNA Single Indexes kits A and B Illumina San Diego CA according to manufacturer's instructions. Briefly cytoplasmic and mitochondrial ribosomal RNA rRNA was removed using biotinylated target specific oligos combined with Ribo Zero rRNA removal beads. Following purification the depleted RNA was fragmented into small pieces using divalent cations at 94oC for 2 minutes. Cleaved RNA fragments were then copied into first strand cDNA using reverse transcriptase and random primers followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. Strand specificity was achieved by replacing dTTP with dUTP during second strand synthesis. The double stranded cDNA fragments were blunted using T4 DNA polymerase Klenow DNA polymerase and T4 PNK. A single 'A' nucleotide was added to the three prime ends of the blunt DNA fragments using a Klenow fragment three prime to five primeexo minus enzyme. The cDNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR amplification 30 sec at 98oC; [10 sec at 98oC 30 sec at 60oC 30 sec at 72oC] x 12 cycles; 5 min at 72oC. Surplus PCR primers were further removed by purification using AMPure XP beads Beckman Coulter Villepinte France and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis.   tissue:embryo|genotype:wild type|phenotype:unaffected epib|Stage:5 hpf GSM3968576 GSM3968576: WT 3; Danio rerio; RNA Seq GSM3968576   1 Embryos were sampled in liquid nitrogen and stored at 80°C until further processing. RNA was extracted with TRIzol Life Technologies following the manufacturer's instructions with minor modifications: 30 larvae were sampled in a volume of 1.5 ml TRIzol reagent. Samples were kept at least overnight at 80°C. In order to disrupt the cell membranes the samples were passed four times through a syringe with a diameter of 0.45 mm. Total RNA Seq libraries were generated from 500 ng of total RNA using TruSeq Stranded Total RNA Library Prep Human/Mouse/Rat kit and TruSeq RNA Single Indexes kits A and B Illumina San Diego CA according to manufacturer's instructions. Briefly cytoplasmic and mitochondrial ribosomal RNA rRNA was removed using biotinylated target specific oligos combined with Ribo Zero rRNA removal beads. Following purification the depleted RNA was fragmented into small pieces using divalent cations at 94oC for 2 minutes. Cleaved RNA fragments were then copied into first strand cDNA using reverse transcriptase and random primers followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. Strand specificity was achieved by replacing dTTP with dUTP during second strand synthesis. The double stranded cDNA fragments were blunted using T4 DNA polymerase Klenow DNA polymerase and T4 PNK. A single 'A' nucleotide was added to the three prime ends of the blunt DNA fragments using a Klenow fragment three prime to five primeexo minus enzyme. The cDNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR amplification 30 sec at 98oC; [10 sec at 98oC 30 sec at 60oC 30 sec at 72oC] x 12 cycles; 5 min at 72oC. Surplus PCR primers were further removed by purification using AMPure XP beads Beckman Coulter Villepinte France and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis. GEO Accession:GSM3968576 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP216221     VRGN28_S2.R1.fastq.gz VRGN28_S2.R2.fastq.gz fastq fastq 8867395800.0 88673958.0 GSM3968576 r1 0:50 1:50 A:2377675533;C:2037155907;G:2034235985;T:2416929022;N:1399353 50 50     2377675533 2037155907 2034235985 2416929022 1399353 SRX6579572 SRS5145283 SRA925866 GEO Naef Lab, SV, EPFL 2 0.89244 0.89367 0.25364 0.25177 0.74562 0.75071 0.48724 0.49306 50 50 B B biological fallback assumption illumina hiseq_era 3prime rrna_depletion trueseq bulk bulk bulk   Switzerland 2019-07-24 Blastula Embryo Embryo Imprecise All anatomical structures
53286 53286 SRR9822879 SRX6579571 SRS5145282 SRP216221 PRJNA556409 RNA seq of mondoa zebrafish mutants GSE134778 Transcriptome Analysis The glucose sensing Mondo pathway regulates expression of metabolic genes in mammals. Here we have revealed an unexpected role of this pathway in vertebrate embryonic development. We characterized Mondo pathway function in the zebrafish and showed that knock down of mondoa impaired the early morphogenetic movement of epib in zebrafish embryos. Expression of nsdhl an enzyme of cholesterol and pregnenolone synthesis was strongly reduced in these embryos. Loss of Nsdhl function likewise impaired epib and led to microtubule cytoskeleton defects in the embryo similar to MondoA loss of function. both epib and microtubule cytoskeleton defects were partially restored by pregnenolone treatment. Gene disruption of mondoa perturbed epib with only partial penetrance likely due to compensatory changes in the expression of cholesterol/steroid metabolism genes. Collectively our results show a novel role for MondoA in the regulation of early vertebrate development connecting glucose cholesterol and steroid hormone metabolism with early embryonic cell movements. Overall design: RNA seq in mondoa zebrafish mutants parent bioproject:PRJNA603464 pubmed:32969791   WT n 2 GSM3968575   source name:total RNA|tissue:embryo|genotype:wild type|phenotype:unaffected epib|Stage:5 hpf WT n 2 Image analysis and base calling were performed using RTA 2.7.3 and bcl2fastq 2.17.1.14. Sequences were aligned to the full reference genome index build with Ensembl annotation to improve accuracy on splices junctions using STAR 2.4.0i A custom Perl script was used to quantify uniquely mapped reads. For each Ensembl gene all exons of the respective annotated protein coding transcripts were considered. Genome build: zv10 Supplementary files format and content: counts mutants.txt: Tab delimited text file contains raw counts. total RNA   Embryos were sampled in liquid nitrogen and stored at 80°C until further processing. RNA was extracted with TRIzol Life Technologies following the manufacturer’s instructions with minor modifications: 30 larvae were sampled in a volume of 1.5 ml TRIzol reagent. Samples were kept at least overnight at 80°C. In order to disrupt the cell membranes the samples were passed four times through a syringe with a diameter of 0.45 mm. Total RNA Seq libraries were generated from 500 ng of total RNA using TruSeq Stranded Total RNA Library Prep Human/Mouse/Rat kit and TruSeq RNA Single Indexes kits A and B Illumina San Diego CA according to manufacturer's instructions. Briefly cytoplasmic and mitochondrial ribosomal RNA rRNA was removed using biotinylated target specific oligos combined with Ribo Zero rRNA removal beads. Following purification the depleted RNA was fragmented into small pieces using divalent cations at 94oC for 2 minutes. Cleaved RNA fragments were then copied into first strand cDNA using reverse transcriptase and random primers followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. Strand specificity was achieved by replacing dTTP with dUTP during second strand synthesis. The double stranded cDNA fragments were blunted using T4 DNA polymerase Klenow DNA polymerase and T4 PNK. A single 'A' nucleotide was added to the three prime ends of the blunt DNA fragments using a Klenow fragment three prime to five primeexo minus enzyme. The cDNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR amplification 30 sec at 98oC; [10 sec at 98oC 30 sec at 60oC 30 sec at 72oC] x 12 cycles; 5 min at 72oC. Surplus PCR primers were further removed by purification using AMPure XP beads Beckman Coulter Villepinte France and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis.   tissue:embryo|genotype:wild type|phenotype:unaffected epib|Stage:5 hpf GSM3968575 GSM3968575: WT 2; Danio rerio; RNA Seq GSM3968575   1 Embryos were sampled in liquid nitrogen and stored at 80°C until further processing. RNA was extracted with TRIzol Life Technologies following the manufacturer's instructions with minor modifications: 30 larvae were sampled in a volume of 1.5 ml TRIzol reagent. Samples were kept at least overnight at 80°C. In order to disrupt the cell membranes the samples were passed four times through a syringe with a diameter of 0.45 mm. Total RNA Seq libraries were generated from 500 ng of total RNA using TruSeq Stranded Total RNA Library Prep Human/Mouse/Rat kit and TruSeq RNA Single Indexes kits A and B Illumina San Diego CA according to manufacturer's instructions. Briefly cytoplasmic and mitochondrial ribosomal RNA rRNA was removed using biotinylated target specific oligos combined with Ribo Zero rRNA removal beads. Following purification the depleted RNA was fragmented into small pieces using divalent cations at 94oC for 2 minutes. Cleaved RNA fragments were then copied into first strand cDNA using reverse transcriptase and random primers followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. Strand specificity was achieved by replacing dTTP with dUTP during second strand synthesis. The double stranded cDNA fragments were blunted using T4 DNA polymerase Klenow DNA polymerase and T4 PNK. A single 'A' nucleotide was added to the three prime ends of the blunt DNA fragments using a Klenow fragment three prime to five primeexo minus enzyme. The cDNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR amplification 30 sec at 98oC; [10 sec at 98oC 30 sec at 60oC 30 sec at 72oC] x 12 cycles; 5 min at 72oC. Surplus PCR primers were further removed by purification using AMPure XP beads Beckman Coulter Villepinte France and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis. GEO Accession:GSM3968575 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP216221     VRGN27_S2.R1.fastq.gz VRGN27_S2.R2.fastq.gz fastq fastq 8463600400.0 84636004.0 GSM3968575 r1 0:50 1:50 A:2302466173;C:1909305079;G:1886583111;T:2363912593;N:1333444 50 50     2302466173 1909305079 1886583111 2363912593 1333444 SRX6579571 SRS5145282 SRA925866 GEO Naef Lab, SV, EPFL 2 0.88572 0.88835 0.29743 0.29437 0.74919 0.75672 0.49434 0.49314 50 50 B B biological fallback assumption illumina hiseq_era 3prime rrna_depletion trueseq bulk bulk bulk   Switzerland 2019-07-24 Blastula Embryo Embryo Imprecise All anatomical structures
53287 53287 SRR9822878 SRX6579570 SRS5145281 SRP216221 PRJNA556409 RNA seq of mondoa zebrafish mutants GSE134778 Transcriptome Analysis The glucose sensing Mondo pathway regulates expression of metabolic genes in mammals. Here we have revealed an unexpected role of this pathway in vertebrate embryonic development. We characterized Mondo pathway function in the zebrafish and showed that knock down of mondoa impaired the early morphogenetic movement of epib in zebrafish embryos. Expression of nsdhl an enzyme of cholesterol and pregnenolone synthesis was strongly reduced in these embryos. Loss of Nsdhl function likewise impaired epib and led to microtubule cytoskeleton defects in the embryo similar to MondoA loss of function. both epib and microtubule cytoskeleton defects were partially restored by pregnenolone treatment. Gene disruption of mondoa perturbed epib with only partial penetrance likely due to compensatory changes in the expression of cholesterol/steroid metabolism genes. Collectively our results show a novel role for MondoA in the regulation of early vertebrate development connecting glucose cholesterol and steroid hormone metabolism with early embryonic cell movements. Overall design: RNA seq in mondoa zebrafish mutants parent bioproject:PRJNA603464 pubmed:32969791   WT n 1 GSM3968574   source name:total RNA|tissue:embryo|genotype:wild type|phenotype:unaffected epib|Stage:5 hpf WT n 1 Image analysis and base calling were performed using RTA 2.7.3 and bcl2fastq 2.17.1.14. Sequences were aligned to the full reference genome index build with Ensembl annotation to improve accuracy on splices junctions using STAR 2.4.0i A custom Perl script was used to quantify uniquely mapped reads. For each Ensembl gene all exons of the respective annotated protein coding transcripts were considered. Genome build: zv10 Supplementary files format and content: counts mutants.txt: Tab delimited text file contains raw counts. total RNA   Embryos were sampled in liquid nitrogen and stored at 80°C until further processing. RNA was extracted with TRIzol Life Technologies following the manufacturer’s instructions with minor modifications: 30 larvae were sampled in a volume of 1.5 ml TRIzol reagent. Samples were kept at least overnight at 80°C. In order to disrupt the cell membranes the samples were passed four times through a syringe with a diameter of 0.45 mm. Total RNA Seq libraries were generated from 500 ng of total RNA using TruSeq Stranded Total RNA Library Prep Human/Mouse/Rat kit and TruSeq RNA Single Indexes kits A and B Illumina San Diego CA according to manufacturer's instructions. Briefly cytoplasmic and mitochondrial ribosomal RNA rRNA was removed using biotinylated target specific oligos combined with Ribo Zero rRNA removal beads. Following purification the depleted RNA was fragmented into small pieces using divalent cations at 94oC for 2 minutes. Cleaved RNA fragments were then copied into first strand cDNA using reverse transcriptase and random primers followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. Strand specificity was achieved by replacing dTTP with dUTP during second strand synthesis. The double stranded cDNA fragments were blunted using T4 DNA polymerase Klenow DNA polymerase and T4 PNK. A single 'A' nucleotide was added to the three prime ends of the blunt DNA fragments using a Klenow fragment three prime to five primeexo minus enzyme. The cDNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR amplification 30 sec at 98oC; [10 sec at 98oC 30 sec at 60oC 30 sec at 72oC] x 12 cycles; 5 min at 72oC. Surplus PCR primers were further removed by purification using AMPure XP beads Beckman Coulter Villepinte France and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis.   tissue:embryo|genotype:wild type|phenotype:unaffected epib|Stage:5 hpf GSM3968574 GSM3968574: WT 1; Danio rerio; RNA Seq GSM3968574   1 Embryos were sampled in liquid nitrogen and stored at 80°C until further processing. RNA was extracted with TRIzol Life Technologies following the manufacturer's instructions with minor modifications: 30 larvae were sampled in a volume of 1.5 ml TRIzol reagent. Samples were kept at least overnight at 80°C. In order to disrupt the cell membranes the samples were passed four times through a syringe with a diameter of 0.45 mm. Total RNA Seq libraries were generated from 500 ng of total RNA using TruSeq Stranded Total RNA Library Prep Human/Mouse/Rat kit and TruSeq RNA Single Indexes kits A and B Illumina San Diego CA according to manufacturer's instructions. Briefly cytoplasmic and mitochondrial ribosomal RNA rRNA was removed using biotinylated target specific oligos combined with Ribo Zero rRNA removal beads. Following purification the depleted RNA was fragmented into small pieces using divalent cations at 94oC for 2 minutes. Cleaved RNA fragments were then copied into first strand cDNA using reverse transcriptase and random primers followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. Strand specificity was achieved by replacing dTTP with dUTP during second strand synthesis. The double stranded cDNA fragments were blunted using T4 DNA polymerase Klenow DNA polymerase and T4 PNK. A single 'A' nucleotide was added to the three prime ends of the blunt DNA fragments using a Klenow fragment three prime to five primeexo minus enzyme. The cDNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR amplification 30 sec at 98oC; [10 sec at 98oC 30 sec at 60oC 30 sec at 72oC] x 12 cycles; 5 min at 72oC. Surplus PCR primers were further removed by purification using AMPure XP beads Beckman Coulter Villepinte France and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis. GEO Accession:GSM3968574 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP216221     VRGN26_S2.R1.fastq.gz VRGN26_S2.R2.fastq.gz fastq fastq 7628784300.0 76287843.0 GSM3968574 r1 0:50 1:50 A:2078754408;C:1719651241;G:1693975000;T:2135219453;N:1184198 50 50     2078754408 1719651241 1693975000 2135219453 1184198 SRX6579570 SRS5145281 SRA925866 GEO Naef Lab, SV, EPFL 2 0.89762 0.89981 0.30018 0.29512 0.74059 0.74846 0.48922 0.49838 50 50 B B biological fallback assumption illumina hiseq_era 3prime rrna_depletion trueseq bulk bulk bulk   Switzerland 2019-07-24 Blastula Embryo Embryo Imprecise All anatomical structures
53288 53288 SRR9822877 SRX6579569 SRS5145280 SRP216221 PRJNA556409 RNA seq of mondoa zebrafish mutants GSE134778 Transcriptome Analysis The glucose sensing Mondo pathway regulates expression of metabolic genes in mammals. Here we have revealed an unexpected role of this pathway in vertebrate embryonic development. We characterized Mondo pathway function in the zebrafish and showed that knock down of mondoa impaired the early morphogenetic movement of epib in zebrafish embryos. Expression of nsdhl an enzyme of cholesterol and pregnenolone synthesis was strongly reduced in these embryos. Loss of Nsdhl function likewise impaired epib and led to microtubule cytoskeleton defects in the embryo similar to MondoA loss of function. both epib and microtubule cytoskeleton defects were partially restored by pregnenolone treatment. Gene disruption of mondoa perturbed epib with only partial penetrance likely due to compensatory changes in the expression of cholesterol/steroid metabolism genes. Collectively our results show a novel role for MondoA in the regulation of early vertebrate development connecting glucose cholesterol and steroid hormone metabolism with early embryonic cell movements. Overall design: RNA seq in mondoa zebrafish mutants parent bioproject:PRJNA603464 pubmed:32969791   mondoa n 3 GSM3968573   source name:total RNA|tissue:embryo|genotype:MZmondoA mutant allel: ka405 ZFIN ID: ZDB ALT 180628 2|phenotype:unaffected epib|Stage:5 hpf mondoa n 3 Image analysis and base calling were performed using RTA 2.7.3 and bcl2fastq 2.17.1.14. Sequences were aligned to the full reference genome index build with Ensembl annotation to improve accuracy on splices junctions using STAR 2.4.0i A custom Perl script was used to quantify uniquely mapped reads. For each Ensembl gene all exons of the respective annotated protein coding transcripts were considered. Genome build: zv10 Supplementary files format and content: counts mutants.txt: Tab delimited text file contains raw counts. total RNA   Embryos were sampled in liquid nitrogen and stored at 80°C until further processing. RNA was extracted with TRIzol Life Technologies following the manufacturer’s instructions with minor modifications: 30 larvae were sampled in a volume of 1.5 ml TRIzol reagent. Samples were kept at least overnight at 80°C. In order to disrupt the cell membranes the samples were passed four times through a syringe with a diameter of 0.45 mm. Total RNA Seq libraries were generated from 500 ng of total RNA using TruSeq Stranded Total RNA Library Prep Human/Mouse/Rat kit and TruSeq RNA Single Indexes kits A and B Illumina San Diego CA according to manufacturer's instructions. Briefly cytoplasmic and mitochondrial ribosomal RNA rRNA was removed using biotinylated target specific oligos combined with Ribo Zero rRNA removal beads. Following purification the depleted RNA was fragmented into small pieces using divalent cations at 94oC for 2 minutes. Cleaved RNA fragments were then copied into first strand cDNA using reverse transcriptase and random primers followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. Strand specificity was achieved by replacing dTTP with dUTP during second strand synthesis. The double stranded cDNA fragments were blunted using T4 DNA polymerase Klenow DNA polymerase and T4 PNK. A single 'A' nucleotide was added to the three prime ends of the blunt DNA fragments using a Klenow fragment three prime to five primeexo minus enzyme. The cDNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR amplification 30 sec at 98oC; [10 sec at 98oC 30 sec at 60oC 30 sec at 72oC] x 12 cycles; 5 min at 72oC. Surplus PCR primers were further removed by purification using AMPure XP beads Beckman Coulter Villepinte France and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis.   tissue:embryo|genotype:MZmondoA mutant allel: ka405 ZFIN ID: ZDB ALT 180628 2|phenotype:unaffected epib|Stage:5 hpf GSM3968573 GSM3968573: mondoa 3; Danio rerio; RNA Seq GSM3968573   1 Embryos were sampled in liquid nitrogen and stored at 80°C until further processing. RNA was extracted with TRIzol Life Technologies following the manufacturer's instructions with minor modifications: 30 larvae were sampled in a volume of 1.5 ml TRIzol reagent. Samples were kept at least overnight at 80°C. In order to disrupt the cell membranes the samples were passed four times through a syringe with a diameter of 0.45 mm. Total RNA Seq libraries were generated from 500 ng of total RNA using TruSeq Stranded Total RNA Library Prep Human/Mouse/Rat kit and TruSeq RNA Single Indexes kits A and B Illumina San Diego CA according to manufacturer's instructions. Briefly cytoplasmic and mitochondrial ribosomal RNA rRNA was removed using biotinylated target specific oligos combined with Ribo Zero rRNA removal beads. Following purification the depleted RNA was fragmented into small pieces using divalent cations at 94oC for 2 minutes. Cleaved RNA fragments were then copied into first strand cDNA using reverse transcriptase and random primers followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. Strand specificity was achieved by replacing dTTP with dUTP during second strand synthesis. The double stranded cDNA fragments were blunted using T4 DNA polymerase Klenow DNA polymerase and T4 PNK. A single 'A' nucleotide was added to the three prime ends of the blunt DNA fragments using a Klenow fragment three prime to five primeexo minus enzyme. The cDNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR amplification 30 sec at 98oC; [10 sec at 98oC 30 sec at 60oC 30 sec at 72oC] x 12 cycles; 5 min at 72oC. Surplus PCR primers were further removed by purification using AMPure XP beads Beckman Coulter Villepinte France and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis. GEO Accession:GSM3968573 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP216221     VRGN25_S2.R2.fastq.gz VRGN25_S2.R1.fastq.gz fastq fastq 7551918900.0 75519189.0 GSM3968573 r1 0:50 1:50 A:2066041718;C:1702385113;G:1686618839;T:2095697396;N:1175834 50 50     2066041718 1702385113 1686618839 2095697396 1175834 SRX6579569 SRS5145280 SRA925866 GEO Naef Lab, SV, EPFL 2 0.89704 0.89919 0.27361 0.27325 0.74637 0.75755 0.50569 0.5107 50 50 B B biological fallback assumption illumina hiseq_era 3prime rrna_depletion trueseq bulk bulk bulk   Switzerland 2019-07-24 Blastula Embryo Embryo Imprecise All anatomical structures
53289 53289 SRR9822876 SRX6579568 SRS5145279 SRP216221 PRJNA556409 RNA seq of mondoa zebrafish mutants GSE134778 Transcriptome Analysis The glucose sensing Mondo pathway regulates expression of metabolic genes in mammals. Here we have revealed an unexpected role of this pathway in vertebrate embryonic development. We characterized Mondo pathway function in the zebrafish and showed that knock down of mondoa impaired the early morphogenetic movement of epib in zebrafish embryos. Expression of nsdhl an enzyme of cholesterol and pregnenolone synthesis was strongly reduced in these embryos. Loss of Nsdhl function likewise impaired epib and led to microtubule cytoskeleton defects in the embryo similar to MondoA loss of function. both epib and microtubule cytoskeleton defects were partially restored by pregnenolone treatment. Gene disruption of mondoa perturbed epib with only partial penetrance likely due to compensatory changes in the expression of cholesterol/steroid metabolism genes. Collectively our results show a novel role for MondoA in the regulation of early vertebrate development connecting glucose cholesterol and steroid hormone metabolism with early embryonic cell movements. Overall design: RNA seq in mondoa zebrafish mutants parent bioproject:PRJNA603464 pubmed:32969791   mondoa n 2 GSM3968572   source name:total RNA|tissue:embryo|genotype:MZmondoA mutant allel: ka405 ZFIN ID: ZDB ALT 180628 2|phenotype:unaffected epib|Stage:5 hpf mondoa n 2 Image analysis and base calling were performed using RTA 2.7.3 and bcl2fastq 2.17.1.14. Sequences were aligned to the full reference genome index build with Ensembl annotation to improve accuracy on splices junctions using STAR 2.4.0i A custom Perl script was used to quantify uniquely mapped reads. For each Ensembl gene all exons of the respective annotated protein coding transcripts were considered. Genome build: zv10 Supplementary files format and content: counts mutants.txt: Tab delimited text file contains raw counts. total RNA   Embryos were sampled in liquid nitrogen and stored at 80°C until further processing. RNA was extracted with TRIzol Life Technologies following the manufacturer’s instructions with minor modifications: 30 larvae were sampled in a volume of 1.5 ml TRIzol reagent. Samples were kept at least overnight at 80°C. In order to disrupt the cell membranes the samples were passed four times through a syringe with a diameter of 0.45 mm. Total RNA Seq libraries were generated from 500 ng of total RNA using TruSeq Stranded Total RNA Library Prep Human/Mouse/Rat kit and TruSeq RNA Single Indexes kits A and B Illumina San Diego CA according to manufacturer's instructions. Briefly cytoplasmic and mitochondrial ribosomal RNA rRNA was removed using biotinylated target specific oligos combined with Ribo Zero rRNA removal beads. Following purification the depleted RNA was fragmented into small pieces using divalent cations at 94oC for 2 minutes. Cleaved RNA fragments were then copied into first strand cDNA using reverse transcriptase and random primers followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. Strand specificity was achieved by replacing dTTP with dUTP during second strand synthesis. The double stranded cDNA fragments were blunted using T4 DNA polymerase Klenow DNA polymerase and T4 PNK. A single 'A' nucleotide was added to the three prime ends of the blunt DNA fragments using a Klenow fragment three prime to five primeexo minus enzyme. The cDNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR amplification 30 sec at 98oC; [10 sec at 98oC 30 sec at 60oC 30 sec at 72oC] x 12 cycles; 5 min at 72oC. Surplus PCR primers were further removed by purification using AMPure XP beads Beckman Coulter Villepinte France and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis.   tissue:embryo|genotype:MZmondoA mutant allel: ka405 ZFIN ID: ZDB ALT 180628 2|phenotype:unaffected epib|Stage:5 hpf GSM3968572 GSM3968572: mondoa 2; Danio rerio; RNA Seq GSM3968572   1 Embryos were sampled in liquid nitrogen and stored at 80°C until further processing. RNA was extracted with TRIzol Life Technologies following the manufacturer's instructions with minor modifications: 30 larvae were sampled in a volume of 1.5 ml TRIzol reagent. Samples were kept at least overnight at 80°C. In order to disrupt the cell membranes the samples were passed four times through a syringe with a diameter of 0.45 mm. Total RNA Seq libraries were generated from 500 ng of total RNA using TruSeq Stranded Total RNA Library Prep Human/Mouse/Rat kit and TruSeq RNA Single Indexes kits A and B Illumina San Diego CA according to manufacturer's instructions. Briefly cytoplasmic and mitochondrial ribosomal RNA rRNA was removed using biotinylated target specific oligos combined with Ribo Zero rRNA removal beads. Following purification the depleted RNA was fragmented into small pieces using divalent cations at 94oC for 2 minutes. Cleaved RNA fragments were then copied into first strand cDNA using reverse transcriptase and random primers followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. Strand specificity was achieved by replacing dTTP with dUTP during second strand synthesis. The double stranded cDNA fragments were blunted using T4 DNA polymerase Klenow DNA polymerase and T4 PNK. A single 'A' nucleotide was added to the three prime ends of the blunt DNA fragments using a Klenow fragment three prime to five primeexo minus enzyme. The cDNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR amplification 30 sec at 98oC; [10 sec at 98oC 30 sec at 60oC 30 sec at 72oC] x 12 cycles; 5 min at 72oC. Surplus PCR primers were further removed by purification using AMPure XP beads Beckman Coulter Villepinte France and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis. GEO Accession:GSM3968572 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP216221     VRGN24_S2.R1.fastq.gz VRGN24_S2.R2.fastq.gz fastq fastq 7666458300.0 76664583.0 GSM3968572 r1 0:50 1:50 A:2108884078;C:1705527841;G:1677597808;T:2173267709;N:1180864 50 50     2108884078 1705527841 1677597808 2173267709 1180864 SRX6579568 SRS5145279 SRA925866 GEO Naef Lab, SV, EPFL 2 0.89316 0.89507 0.29969 0.29911 0.74947 0.75741 0.50817 0.50943 50 50 B B biological fallback assumption illumina hiseq_era 3prime rrna_depletion trueseq bulk bulk bulk   Switzerland 2019-07-24 Blastula Embryo Embryo Imprecise All anatomical structures
53290 53290 SRR9822875 SRX6579567 SRS5145278 SRP216221 PRJNA556409 RNA seq of mondoa zebrafish mutants GSE134778 Transcriptome Analysis The glucose sensing Mondo pathway regulates expression of metabolic genes in mammals. Here we have revealed an unexpected role of this pathway in vertebrate embryonic development. We characterized Mondo pathway function in the zebrafish and showed that knock down of mondoa impaired the early morphogenetic movement of epib in zebrafish embryos. Expression of nsdhl an enzyme of cholesterol and pregnenolone synthesis was strongly reduced in these embryos. Loss of Nsdhl function likewise impaired epib and led to microtubule cytoskeleton defects in the embryo similar to MondoA loss of function. both epib and microtubule cytoskeleton defects were partially restored by pregnenolone treatment. Gene disruption of mondoa perturbed epib with only partial penetrance likely due to compensatory changes in the expression of cholesterol/steroid metabolism genes. Collectively our results show a novel role for MondoA in the regulation of early vertebrate development connecting glucose cholesterol and steroid hormone metabolism with early embryonic cell movements. Overall design: RNA seq in mondoa zebrafish mutants parent bioproject:PRJNA603464 pubmed:32969791   mondoa n 1 GSM3968571   source name:total RNA|tissue:embryo|genotype:MZmondoA mutant allel: ka405 ZFIN ID: ZDB ALT 180628 2|phenotype:unaffected epib|Stage:5 hpf mondoa n 1 Image analysis and base calling were performed using RTA 2.7.3 and bcl2fastq 2.17.1.14. Sequences were aligned to the full reference genome index build with Ensembl annotation to improve accuracy on splices junctions using STAR 2.4.0i A custom Perl script was used to quantify uniquely mapped reads. For each Ensembl gene all exons of the respective annotated protein coding transcripts were considered. Genome build: zv10 Supplementary files format and content: counts mutants.txt: Tab delimited text file contains raw counts. total RNA   Embryos were sampled in liquid nitrogen and stored at 80°C until further processing. RNA was extracted with TRIzol Life Technologies following the manufacturer’s instructions with minor modifications: 30 larvae were sampled in a volume of 1.5 ml TRIzol reagent. Samples were kept at least overnight at 80°C. In order to disrupt the cell membranes the samples were passed four times through a syringe with a diameter of 0.45 mm. Total RNA Seq libraries were generated from 500 ng of total RNA using TruSeq Stranded Total RNA Library Prep Human/Mouse/Rat kit and TruSeq RNA Single Indexes kits A and B Illumina San Diego CA according to manufacturer's instructions. Briefly cytoplasmic and mitochondrial ribosomal RNA rRNA was removed using biotinylated target specific oligos combined with Ribo Zero rRNA removal beads. Following purification the depleted RNA was fragmented into small pieces using divalent cations at 94oC for 2 minutes. Cleaved RNA fragments were then copied into first strand cDNA using reverse transcriptase and random primers followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. Strand specificity was achieved by replacing dTTP with dUTP during second strand synthesis. The double stranded cDNA fragments were blunted using T4 DNA polymerase Klenow DNA polymerase and T4 PNK. A single 'A' nucleotide was added to the three prime ends of the blunt DNA fragments using a Klenow fragment three prime to five primeexo minus enzyme. The cDNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR amplification 30 sec at 98oC; [10 sec at 98oC 30 sec at 60oC 30 sec at 72oC] x 12 cycles; 5 min at 72oC. Surplus PCR primers were further removed by purification using AMPure XP beads Beckman Coulter Villepinte France and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis.   tissue:embryo|genotype:MZmondoA mutant allel: ka405 ZFIN ID: ZDB ALT 180628 2|phenotype:unaffected epib|Stage:5 hpf GSM3968571 GSM3968571: mondoa 1; Danio rerio; RNA Seq GSM3968571   1 Embryos were sampled in liquid nitrogen and stored at 80°C until further processing. RNA was extracted with TRIzol Life Technologies following the manufacturer's instructions with minor modifications: 30 larvae were sampled in a volume of 1.5 ml TRIzol reagent. Samples were kept at least overnight at 80°C. In order to disrupt the cell membranes the samples were passed four times through a syringe with a diameter of 0.45 mm. Total RNA Seq libraries were generated from 500 ng of total RNA using TruSeq Stranded Total RNA Library Prep Human/Mouse/Rat kit and TruSeq RNA Single Indexes kits A and B Illumina San Diego CA according to manufacturer's instructions. Briefly cytoplasmic and mitochondrial ribosomal RNA rRNA was removed using biotinylated target specific oligos combined with Ribo Zero rRNA removal beads. Following purification the depleted RNA was fragmented into small pieces using divalent cations at 94oC for 2 minutes. Cleaved RNA fragments were then copied into first strand cDNA using reverse transcriptase and random primers followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. Strand specificity was achieved by replacing dTTP with dUTP during second strand synthesis. The double stranded cDNA fragments were blunted using T4 DNA polymerase Klenow DNA polymerase and T4 PNK. A single 'A' nucleotide was added to the three prime ends of the blunt DNA fragments using a Klenow fragment three prime to five primeexo minus enzyme. The cDNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR amplification 30 sec at 98oC; [10 sec at 98oC 30 sec at 60oC 30 sec at 72oC] x 12 cycles; 5 min at 72oC. Surplus PCR primers were further removed by purification using AMPure XP beads Beckman Coulter Villepinte France and the final cDNA libraries were checked for quality and quantified using capillary electrophoresis. GEO Accession:GSM3968571 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP216221     VRGN23_S2.R2.fastq.gz VRGN23_S2.R1.fastq.gz fastq fastq 6872302500.0 68723025.0 GSM3968571 r1 0:50 1:50 A:1873323778;C:1557060731;G:1546029001;T:1894832528;N:1056462 50 50     1873323778 1557060731 1546029001 1894832528 1056462 SRX6579567 SRS5145278 SRA925866 GEO Naef Lab, SV, EPFL 2 0.89971 0.90355 0.27138 0.27147 0.7501 0.76185 0.50874 0.51203 50 50 B B biological fallback assumption illumina hiseq_era 3prime rrna_depletion trueseq bulk bulk bulk   Switzerland 2019-07-24 Blastula Embryo Embryo Imprecise All anatomical structures
56025 56025 SRR10836792 SRX7508634 SRS5949026 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   Dome Cyt GSM4252155   source name:Dome stage embryo|tissue:zebrafish embryos|developmental stage:Dome|fraction:Cytosolic Dome Cyt RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages Dome stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:Dome|fraction:Cytosolic GSM4252155 GSM4252155: Dome Cyt; Danio rerio; RNA Seq GSM4252155   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252155 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     Dome-C2_L001_R2_001.fastq.gz Dome-C2_L001_R1_001.fastq.gz fastq fastq 1600183273.0 10609142.0 GSM4252155 r1 0:75.47 1:75.36 A:406319131;C:381620266;G:406104578;T:405721915;N:417383 75 75     406319131 381620266 406104578 405721915 417383 SRX7508634 SRS5949026 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.88183 0.88249 0.10573 0.10435 0.75481 0.76033 0.65717 0.6586 76 75 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56026 56026 SRR10836793 SRX7508634 SRS5949026 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   Dome Cyt GSM4252155   source name:Dome stage embryo|tissue:zebrafish embryos|developmental stage:Dome|fraction:Cytosolic Dome Cyt RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages Dome stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:Dome|fraction:Cytosolic GSM4252155 GSM4252155: Dome Cyt; Danio rerio; RNA Seq GSM4252155   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252155 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     Dome-C2_L002_R1_001.fastq.gz Dome-C2_L002_R2_001.fastq.gz fastq fastq 1566311827.0 10384368.0 GSM4252155 r2 0:75.48 1:75.36 A:398043893;C:372803129;G:397876607;T:397120910;N:467288 75 75     398043893 372803129 397876607 397120910 467288 SRX7508634 SRS5949026 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.8823 0.88437 0.10602 0.10527 0.75564 0.76118 0.65836 0.66025 75 75 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56027 56027 SRR10836794 SRX7508634 SRS5949026 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   Dome Cyt GSM4252155   source name:Dome stage embryo|tissue:zebrafish embryos|developmental stage:Dome|fraction:Cytosolic Dome Cyt RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages Dome stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:Dome|fraction:Cytosolic GSM4252155 GSM4252155: Dome Cyt; Danio rerio; RNA Seq GSM4252155   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252155 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     Dome-C2_L003_R1_001.fastq.gz Dome-C2_L003_R2_001.fastq.gz fastq fastq 1594543994.0 10571557.0 GSM4252155 r3 0:75.47 1:75.36 A:405942855;C:382158076;G:399619556;T:406535479;N:288028 75 75     405942855 382158076 399619556 406535479 288028 SRX7508634 SRS5949026 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.88258 0.88139 0.10614 0.10462 0.754 0.75919 0.3912 0.65984 73 75 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56028 56028 SRR10836795 SRX7508634 SRS5949026 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   Dome Cyt GSM4252155   source name:Dome stage embryo|tissue:zebrafish embryos|developmental stage:Dome|fraction:Cytosolic Dome Cyt RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages Dome stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:Dome|fraction:Cytosolic GSM4252155 GSM4252155: Dome Cyt; Danio rerio; RNA Seq GSM4252155   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252155 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     Dome-C2_L004_R2_001.fastq.gz Dome-C2_L004_R1_001.fastq.gz fastq fastq 1561211427.0 10350323.0 GSM4252155 r4 0:75.48 1:75.36 A:397911507;C:374246496;G:390504606;T:398280725;N:268093 75 75     397911507 374246496 390504606 398280725 268093 SRX7508634 SRS5949026 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.88303 0.88191 0.1066 0.10452 0.75592 0.76 0.65797 0.65812 76 76 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56029 56029 SRR10836788 SRX7508633 SRS5949025 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   Dome Nuc GSM4252154   source name:Dome stage embryo|tissue:zebrafish embryos|developmental stage:Dome|fraction:Nuclear Dome Nuc RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages Dome stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:Dome|fraction:Nuclear GSM4252154 GSM4252154: Dome Nuc; Danio rerio; RNA Seq GSM4252154   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252154 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     Dome-N2_L001_R1_001.fastq.gz Dome-N2_L001_R2_001.fastq.gz fastq fastq 2074718856.0 13753362.0 GSM4252154 r1 0:75.47 1:75.38 A:548207869;C:474119517;G:498330967;T:553614124;N:446379 75 75     548207869 474119517 498330967 553614124 446379 SRX7508633 SRS5949025 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.92904 0.93027 0.25794 0.25635 0.73314 0.73543 0.63799 0.63772 76 75 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56030 56030 SRR10836789 SRX7508633 SRS5949025 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   Dome Nuc GSM4252154   source name:Dome stage embryo|tissue:zebrafish embryos|developmental stage:Dome|fraction:Nuclear Dome Nuc RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages Dome stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:Dome|fraction:Nuclear GSM4252154 GSM4252154: Dome Nuc; Danio rerio; RNA Seq GSM4252154   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252154 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     Dome-N2_L002_R1_001.fastq.gz Dome-N2_L002_R2_001.fastq.gz fastq fastq 2029358370.0 13452695.0 GSM4252154 r2 0:75.47 1:75.38 A:536416032;C:463133536;G:487927693;T:541364778;N:516331 75 75     536416032 463133536 487927693 541364778 516331 SRX7508633 SRS5949025 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.92826 0.93051 0.25765 0.25606 0.73294 0.73537 0.63631 0.63767 75 76 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56031 56031 SRR10836790 SRX7508633 SRS5949025 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   Dome Nuc GSM4252154   source name:Dome stage embryo|tissue:zebrafish embryos|developmental stage:Dome|fraction:Nuclear Dome Nuc RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages Dome stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:Dome|fraction:Nuclear GSM4252154 GSM4252154: Dome Nuc; Danio rerio; RNA Seq GSM4252154   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252154 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     Dome-N2_L003_R1_001.fastq.gz Dome-N2_L003_R2_001.fastq.gz fastq fastq 2071038727.0 13728438.0 GSM4252154 r3 0:75.47 1:75.38 A:549024994;C:475436351;G:490887464;T:555409952;N:279966 75 75     549024994 475436351 490887464 555409952 279966 SRX7508633 SRS5949025 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.92832 0.92953 0.25562 0.25507 0.73253 0.73523 0.64216 0.64054 76 75 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56032 56032 SRR10836791 SRX7508633 SRS5949025 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   Dome Nuc GSM4252154   source name:Dome stage embryo|tissue:zebrafish embryos|developmental stage:Dome|fraction:Nuclear Dome Nuc RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages Dome stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:Dome|fraction:Nuclear GSM4252154 GSM4252154: Dome Nuc; Danio rerio; RNA Seq GSM4252154   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252154 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     Dome-N2_L004_R1_001.fastq.gz Dome-N2_L004_R2_001.fastq.gz fastq fastq 2029324602.0 13451857.0 GSM4252154 r4 0:75.47 1:75.38 A:538432870;C:466166947;G:479781700;T:544692254;N:250831 75 75     538432870 466166947 479781700 544692254 250831 SRX7508633 SRS5949025 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.92876 0.93022 0.25694 0.25536 0.73186 0.73501 0.64101 0.64 74 73 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56033 56033 SRR10836784 SRX7508632 SRS5949024 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   1000 cell Cyt GSM4252153   source name:1000Cell stage embryo|tissue:zebrafish embryos|developmental stage:1000 cell|fraction:Cytosolic 1000 cell Cyt RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages 1000Cell stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:1000 cell|fraction:Cytosolic GSM4252153 GSM4252153: 1000 cell Cyt; Danio rerio; RNA Seq GSM4252153   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252153 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     1000-Cell-C2_L001_R1_001.fastq.gz 1000-Cell-C2_L001_R2_001.fastq.gz fastq fastq 1502587165.0 9955261.0 GSM4252153 r1 0:75.54 1:75.39 A:375894288;C:354892799;G:392076871;T:379250340;N:472867 75 75     375894288 354892799 392076871 379250340 472867 SRX7508632 SRS5949024 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.95863 0.95851 0.05633 0.05543 0.76171 0.76495 0.61191 0.62248 76 76 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56034 56034 SRR10836785 SRX7508632 SRS5949024 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   1000 cell Cyt GSM4252153   source name:1000Cell stage embryo|tissue:zebrafish embryos|developmental stage:1000 cell|fraction:Cytosolic 1000 cell Cyt RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages 1000Cell stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:1000 cell|fraction:Cytosolic GSM4252153 GSM4252153: 1000 cell Cyt; Danio rerio; RNA Seq GSM4252153   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252153 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     1000-Cell-C2_L002_R1_001.fastq.gz 1000-Cell-C2_L002_R2_001.fastq.gz fastq fastq 1457310394.0 9656279.0 GSM4252153 r2 0:75.53 1:75.38 A:368230607;C:346079382;G:372752678;T:369738027;N:509700 75 75     368230607 346079382 372752678 369738027 509700 SRX7508632 SRS5949024 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.96005 0.95978 0.05635 0.05584 0.76181 0.76449 0.61792 0.61931 71 75 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56035 56035 SRR10836786 SRX7508632 SRS5949024 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   1000 cell Cyt GSM4252153   source name:1000Cell stage embryo|tissue:zebrafish embryos|developmental stage:1000 cell|fraction:Cytosolic 1000 cell Cyt RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages 1000Cell stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:1000 cell|fraction:Cytosolic GSM4252153 GSM4252153: 1000 cell Cyt; Danio rerio; RNA Seq GSM4252153   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252153 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     1000-Cell-C2_L003_R1_001.fastq.gz 1000-Cell-C2_L003_R2_001.fastq.gz fastq fastq 1483084690.0 9826007.0 GSM4252153 r3 0:75.54 1:75.40 A:373953592;C:352136477;G:380681829;T:376135696;N:177096 75 75     373953592 352136477 380681829 376135696 177096 SRX7508632 SRS5949024 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.95919 0.95984 0.05678 0.05526 0.7611 0.76366 0.62126 0.62066 76 75 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56036 56036 SRR10836787 SRX7508632 SRS5949024 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   1000 cell Cyt GSM4252153   source name:1000Cell stage embryo|tissue:zebrafish embryos|developmental stage:1000 cell|fraction:Cytosolic 1000 cell Cyt RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages 1000Cell stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:1000 cell|fraction:Cytosolic GSM4252153 GSM4252153: 1000 cell Cyt; Danio rerio; RNA Seq GSM4252153   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252153 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     1000-Cell-C2_L004_R1_001.fastq.gz 1000-Cell-C2_L004_R2_001.fastq.gz fastq fastq 1429141217.0 9469098.0 GSM4252153 r4 0:75.53 1:75.40 A:363231227;C:342106867;G:358226946;T:365368980;N:207197 75 75     363231227 342106867 358226946 365368980 207197 SRX7508632 SRS5949024 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.95926 0.95888 0.05715 0.05517 0.76096 0.76469 0.62196 0.39902 76 76 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56037 56037 SRR10836780 SRX7508631 SRS5949023 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   1000 cell Nuc GSM4252152   source name:1000Cell stage embryo|tissue:zebrafish embryos|developmental stage:1000 cell|fraction:Nuclear 1000 cell Nuc RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages 1000Cell stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:1000 cell|fraction:Nuclear GSM4252152 GSM4252152: 1000 cell Nuc; Danio rerio; RNA Seq GSM4252152   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252152 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     1000-Cell-N2_L001_R2_001.fastq.gz 1000-Cell-N2_L001_R1_001.fastq.gz fastq fastq 1595563955.0 10573494.0 GSM4252152 r1 0:75.54 1:75.36 A:421378481;C:358086779;G:389857864;T:425749482;N:491349 75 75     421378481 358086779 389857864 425749482 491349 SRX7508631 SRS5949023 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.95517 0.95344 0.06181 0.06032 0.77646 0.7807 0.76076 0.75765 75 76 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56038 56038 SRR10836781 SRX7508631 SRS5949023 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   1000 cell Nuc GSM4252152   source name:1000Cell stage embryo|tissue:zebrafish embryos|developmental stage:1000 cell|fraction:Nuclear 1000 cell Nuc RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages 1000Cell stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:1000 cell|fraction:Nuclear GSM4252152 GSM4252152: 1000 cell Nuc; Danio rerio; RNA Seq GSM4252152   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252152 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     1000-Cell-N2_L002_R1_001.fastq.gz 1000-Cell-N2_L002_R2_001.fastq.gz fastq fastq 1548947081.0 10265666.0 GSM4252152 r2 0:75.54 1:75.35 A:412283151;C:349200776;G:371851315;T:415086710;N:525129 75 75     412283151 349200776 371851315 415086710 525129 SRX7508631 SRS5949023 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.95432 0.9535 0.06288 0.06218 0.77617 0.78027 0.75937 0.75829 75 76 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56039 56039 SRR10836782 SRX7508631 SRS5949023 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   1000 cell Nuc GSM4252152   source name:1000Cell stage embryo|tissue:zebrafish embryos|developmental stage:1000 cell|fraction:Nuclear 1000 cell Nuc RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages 1000Cell stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:1000 cell|fraction:Nuclear GSM4252152 GSM4252152: 1000 cell Nuc; Danio rerio; RNA Seq GSM4252152   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252152 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     1000-Cell-N2_L003_R1_001.fastq.gz 1000-Cell-N2_L003_R2_001.fastq.gz fastq fastq 1577107128.0 10451104.0 GSM4252152 r3 0:75.54 1:75.36 A:419102048;C:355504687;G:379988890;T:422345890;N:165613 75 75     419102048 355504687 379988890 422345890 165613 SRX7508631 SRS5949023 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.95405 0.95325 0.06249 0.06213 0.7766 0.78027 0.76255 0.76017 76 76 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56040 56040 SRR10836783 SRX7508631 SRS5949023 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   1000 cell Nuc GSM4252152   source name:1000Cell stage embryo|tissue:zebrafish embryos|developmental stage:1000 cell|fraction:Nuclear 1000 cell Nuc RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages 1000Cell stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:1000 cell|fraction:Nuclear GSM4252152 GSM4252152: 1000 cell Nuc; Danio rerio; RNA Seq GSM4252152   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252152 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     1000-Cell-N2_L004_R1_001.fastq.gz 1000-Cell-N2_L004_R2_001.fastq.gz fastq fastq 1529237226.0 10134567.0 GSM4252152 r4 0:75.53 1:75.36 A:409232864;C:346856483;G:360903103;T:412045187;N:199589 75 75     409232864 346856483 360903103 412045187 199589 SRX7508631 SRS5949023 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.95294 0.95308 0.06214 0.06138 0.77518 0.77883 0.76284 0.76063 76 76 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56041 56041 SRR10836776 SRX7508630 SRS5949022 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   256 cell Cyt GSM4252151   source name:256Cell stage embryo|tissue:zebrafish embryos|developmental stage:256 cell|fraction:Cytosolic 256 cell Cyt RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages 256Cell stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:256 cell|fraction:Cytosolic GSM4252151 GSM4252151: 256 cell Cyt; Danio rerio; RNA Seq GSM4252151   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252151 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     256-Cell-C2_L001_R1_001.fastq.gz 256-Cell-C2_L001_R2_001.fastq.gz fastq fastq 1366272364.0 9052072.0 GSM4252151 r1 0:75.51 1:75.42 A:323814430;C:338855107;G:375548122;T:327579283;N:475422 75 75     323814430 338855107 375548122 327579283 475422 SRX7508630 SRS5949022 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.83802 0.8313 0.07237 0.06855 0.77352 0.77611 0.60128 0.59485 76 76 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56042 56042 SRR10836777 SRX7508630 SRS5949022 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   256 cell Cyt GSM4252151   source name:256Cell stage embryo|tissue:zebrafish embryos|developmental stage:256 cell|fraction:Cytosolic 256 cell Cyt RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages 256Cell stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:256 cell|fraction:Cytosolic GSM4252151 GSM4252151: 256 cell Cyt; Danio rerio; RNA Seq GSM4252151   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252151 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     256-Cell-C2_L002_R1_001.fastq.gz 256-Cell-C2_L002_R2_001.fastq.gz fastq fastq 1327435639.0 8795964.0 GSM4252151 r2 0:75.50 1:75.41 A:318150528;C:331260745;G:357195153;T:320310778;N:518435 75 75     318150528 331260745 357195153 320310778 518435 SRX7508630 SRS5949022 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.83323 0.82627 0.07204 0.06736 0.77169 0.77437 0.60689 0.5972 76 76 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56043 56043 SRR10836778 SRX7508630 SRS5949022 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   256 cell Cyt GSM4252151   source name:256Cell stage embryo|tissue:zebrafish embryos|developmental stage:256 cell|fraction:Cytosolic 256 cell Cyt RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages 256Cell stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:256 cell|fraction:Cytosolic GSM4252151 GSM4252151: 256 cell Cyt; Danio rerio; RNA Seq GSM4252151   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252151 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     256-Cell-C2_L003_R1_001.fastq.gz 256-Cell-C2_L003_R2_001.fastq.gz fastq fastq 1349181703.0 8939006.0 GSM4252151 r3 0:75.51 1:75.43 A:322588049;C:336503258;G:364782594;T:325107660;N:200142 75 75     322588049 336503258 364782594 325107660 200142 SRX7508630 SRS5949022 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.83327 0.82509 0.07273 0.06804 0.7736 0.77597 0.61092 0.59732 76 76 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56044 56044 SRR10836779 SRX7508630 SRS5949022 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   256 cell Cyt GSM4252151   source name:256Cell stage embryo|tissue:zebrafish embryos|developmental stage:256 cell|fraction:Cytosolic 256 cell Cyt RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages 256Cell stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:256 cell|fraction:Cytosolic GSM4252151 GSM4252151: 256 cell Cyt; Danio rerio; RNA Seq GSM4252151   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252151 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     256-Cell-C2_L004_R1_001.fastq.gz 256-Cell-C2_L004_R2_001.fastq.gz fastq fastq 1303623813.0 8637679.0 GSM4252151 r4 0:75.50 1:75.42 A:314181485;C:328280077;G:344028018;T:316907421;N:226812 75 75     314181485 328280077 344028018 316907421 226812 SRX7508630 SRS5949022 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.82898 0.82179 0.07137 0.06711 0.77177 0.775 0.60119 0.59652 75 75 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56045 56045 SRR10836772 SRX7508629 SRS5949021 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   256 cell Nuc GSM4252150   source name:256Cell stage embryo|tissue:zebrafish embryos|developmental stage:256 cell|fraction:Nuclear 256 cell Nuc RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages 256Cell stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:256 cell|fraction:Nuclear GSM4252150 GSM4252150: 256 cell Nuc; Danio rerio; RNA Seq GSM4252150   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252150 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     256-Cell-N2_L001_R2_001.fastq.gz 256-Cell-N2_L001_R1_001.fastq.gz fastq fastq 1628475646.0 10788030.0 GSM4252150 r1 0:75.57 1:75.38 A:425213330;C:378434245;G:395256706;T:429054367;N:516998 75 75     425213330 378434245 395256706 429054367 516998 SRX7508629 SRS5949021 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.91766 0.91401 0.04416 0.04189 0.79849 0.79961 0.80261 0.79877 76 75 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56046 56046 SRR10836773 SRX7508629 SRS5949021 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   256 cell Nuc GSM4252150   source name:256Cell stage embryo|tissue:zebrafish embryos|developmental stage:256 cell|fraction:Nuclear 256 cell Nuc RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages 256Cell stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:256 cell|fraction:Nuclear GSM4252150 GSM4252150: 256 cell Nuc; Danio rerio; RNA Seq GSM4252150   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252150 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     256-Cell-N2_L002_R1_001.fastq.gz 256-Cell-N2_L002_R2_001.fastq.gz fastq fastq 1582693274.0 10485714.0 GSM4252150 r2 0:75.56 1:75.38 A:415594410;C:368965826;G:379347535;T:418234862;N:550641 75 75     415594410 368965826 379347535 418234862 550641 SRX7508629 SRS5949021 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.91631 0.91267 0.04442 0.04195 0.79638 0.79853 0.8003 0.79303 76 76 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56047 56047 SRR10836774 SRX7508629 SRS5949021 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   256 cell Nuc GSM4252150   source name:256Cell stage embryo|tissue:zebrafish embryos|developmental stage:256 cell|fraction:Nuclear 256 cell Nuc RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages 256Cell stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:256 cell|fraction:Nuclear GSM4252150 GSM4252150: 256 cell Nuc; Danio rerio; RNA Seq GSM4252150   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252150 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     256-Cell-N2_L003_R1_001.fastq.gz 256-Cell-N2_L003_R2_001.fastq.gz fastq fastq 1606644595.0 10643266.0 GSM4252150 r3 0:75.57 1:75.39 A:421472450;C:374504298;G:386000123;T:424481255;N:186469 75 75     421472450 374504298 386000123 424481255 186469 SRX7508629 SRS5949021 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.9166 0.91216 0.04398 0.04143 0.79817 0.79969 0.8036 0.25479 76 76 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
56048 56048 SRR10836775 SRX7508629 SRS5949021 SRP239626 PRJNA599208 Genome wide localization of RNAs during early zebrafish development GSE143208 Transcriptome Analysis We employed transcriptomics methods to examine RNAs from the nuclear and cytosolic fractions from zebrafish embryos at different stages of development. The RNA seq revealed spatial and temporal regulation of lncRNA expression during zebrafish development. Overall design: Localization of RNAs in the nuclear and cytosolic fractions of zebrafish embryonic cells in the 64 Cell 256 Cell 1000 Cell Dome and Shield stages of development.   pubmed:33795334   256 cell Nuc GSM4252150   source name:256Cell stage embryo|tissue:zebrafish embryos|developmental stage:256 cell|fraction:Nuclear 256 cell Nuc RNA seq reads were trimmed using the Trimmomatic v0.27 and then mapped to the Zv9 genome using STAR v2.6.0a with parameters p8 Transcript per million TPM was calculated using the Stringtie and Ballgown program Genome build: Zv9 Supplementary files format and content: tab delimited TPM files for all stages 256Cell stage embryo Dechorionated embryos were then washed twice with 1 X PBS Phosphate buffer solution ThermoFisher. RLN buffer was added to the tube containing embryos and the yolk sac disrupted releasing the cells in the buffer. This was then incubated on ice for 5 mins and centrifuged. The resulting supernatant was collected and labelled as the cytosolic fraction. The pellet was washed twice with 200 µL of RLN buffer and finally collected and labelled as the nuclear fraction. RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. Wild type male and female zebrafish AB strain were set up in breeding tanks overnight and the eggs were collected the next day as soon as 10 min they were laid important to get synchronised embryos. About 100 500 depending upon the stage embryos were collected for each developmental stage 64 cell 256 cell 1000 cell shield 24hpf 50hpf. The embryos were treated with Pronase protease from Streptomyces griseus Sigma Aldrich to remove the chorion 1mL of Pronase working solution 1mg/mL was added to 2 3 ml of fish water containing embryos. tissue:zebrafish embryos|developmental stage:256 cell|fraction:Nuclear GSM4252150 GSM4252150: 256 cell Nuc; Danio rerio; RNA Seq GSM4252150   1 RNA was extracted using the Rneasy mini kit Sequencing libraries were prepared using the TruSeq Stranded Total RNA library Prep kit with the Illumina Ribo Zero rRNA removal kit Human Mouse Rat. GEO Accession:GSM4252150 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP239626     256-Cell-N2_L004_R1_001.fastq.gz 256-Cell-N2_L004_R2_001.fastq.gz fastq fastq 1556236261.0 10309792.0 GSM4252150 r4 0:75.56 1:75.39 A:410297368;C:364669857;G:367737181;T:413315089;N:216766 75 75     410297368 364669857 367737181 413315089 216766 SRX7508629 SRS5949021 SRA1020776 GEO School of Biosciences, University of Birmingham 2 0.91475 0.91068 0.04464 0.04249 0.7974 0.79878 0.80334 0.7997 76 75 B B biological fallback assumption illumina nextseq unknown rrna_depletion ribozero bulk bulk bulk   United Kingdom 2020-01-06 Blastula Embryo Embryo Imprecise All anatomical structures
57241 57241 SRR11293510 SRX7899072 SRS6307327 SRP252498 PRJNA612134 Targeted RNA Knockdown by a Type III CRISPR Cas Complex in Zebrafish GSE146852 Transcriptome Analysis RNA interference RNAi is a powerful experimental tool for RNA knockdown but not all organisms are amenable. Here we provide a “proof of principle” demonstration that CRISPR endoribonuclease can be used for programmable mRNA transcript degradation in eukaryotes. Using zebrafish as the animal model and Csm complex as the CRISPR endoribonuclease we targeted EGFP transcript expressed from a variety of promoters. A drastic decrease of fluorescence was achieved in germ cells of the vasa:EGFP line. Weaker effects were also seen in fish lines that express EGFP zygotically. Knockdown was statistically significant in cmcl2:EGFP and fli1:EGFP zebrafish lines at 1 dpf dpf but reduced to background levels at 2 dpf. The nkx2.5:EGFP fish line was least susceptible to Csm mediated EGFP knockdown. We also tested Csm mediated knockdown on the endogenous tdgf1 oep transcript. At optimal Csm dose we observed a penetrance of the characteristic one eyed phenotype at greater than 50% penetrance and hence with similar efficiency to morpholino mediated knockdown. We conclude that Csm mediated knockdown is very efficient for maternal transcripts and can also be used for mixed maternal/early zygotic and early zygotic transcripts in some cases reaching comparable efficiency to morpholino based knockdown with no significant off target effects in our model. Overall design: Zebrafish embryos from the transgenic line Tgddx4:ddx4 EGFP on the background of ABTL were injected with the Csm complex targeted against the EGFP transcript or mock solution StCsm buffer without xxx proteins. The RNA was isolated from the pooled embryos at three time points: 128 cell stage 5 hpf and 24 hpf. Three replicates were obtained from each experimental condition and time point. Pair end sequencing was performed on the Illumina NextSeq platform.   pubmed:32833532   Mock 5hpf Rep 3 GSM4407869   source name:whole zebrafish embryo|strain:ABTL Tgddx4:ddx4 EGFP|tissue:whole embryo|condition:Mock|developmental stage:5 hpf Mock 5hpf Rep 3 Base calling was performed by Illumina RTA 1.18.61. Files were translated into demultiplexed FASTQ files by Illumina bcl2fastq2 2.16.10. The first 12 nucleotides of read 1 containing the UMI sequence were extracted and added to the sequence identifier using UMI tools v. 1.0.0. Low quality sequences and adapter content were trimmed with cutadapt v. 2.4. Reads were aligned to the GRCz11 reference genome supplemented with the EGFP sequence with STAR v. 2.7.2b using the following custom parameters: outFilterType BySJout outFilterMultimapNmax 20 alignSJoverhangMin 8 alignSJDBoverhangMin 1 outFilterMismatchNmax 999 alignIntronMin 20 alignIntronMax 1000000 alignMatesGapMax 1000000 outFilterMismatchNoverLmax 0.6. Alignments were de duplicated based on their Unique Molecular Identifier sequence using UMI tools v. 1.0.0. De duplicated alignments were counted per exon on the gene level with the htseq count script from the HTSeq v. 0.11.2 package using Ensembl GRCz11 version 97 genome annotation. Further analysis was performed in R v. 3.6.1 programming language. Differential expression was analyzed using DESeq2 v. 1.24.0 package. Genome build: GRCz11 Supplementary files format and content: HTSeq counts.tsv files were generated with htseq count script they contain counts for each gene for each sample in tab delimited format; normalized counts files were generated with DESeq2 they contain library size normalized read counts for each gene for every sample from each time point in tab delimited format; egfp.fa file contains the EGFP sequence in FASTA format; egfp.gtf file contains the custom EGFP annotation in GTF format; egfp crrna.fa file contains the crRNA sequence against the EGFP transcript used in the experiment in FASTA format. whole zebrafish embryo Zebrafish embryos from three different mating pairs of Tgddx4:ddx4 EGFP females with casper males were pooled during collection. D16A StCsmEGFP was diluted with water for injection 1:1 v/v up to a concentration of 0.7mg/ml. For mock injection Tris storage buffer 10mM Tris HCl pH8.5 300mM Nacl 1mM DTT 0.1mM EDTA 50% glycerol was also diluted in the similar manner. 1nl of either StCsm EGFP solution 0.7ng or buffer alone were injected into the yolk of embryos at the 1 cell stage. In total three independent injections were performed. 25 embryos for both StCsm and mock injected per replica set per time point were frozen in liquid nitrogen stored at xxx℃ and processed together. 25 embryos each for both StCsm and mock injected were pooled at xxx cell stage 5 hpf and 24 hpf from xxx biological replicates. Total RNA was extracted from the whole embryos with Tri Reagent Solution Invitrogen according to the manufacturer’s protocol. RNA was further purified using Zymo RNA Clean and Concentrator 5 kit according to manufacturer's protocol. 1.5 ug of total RNA was subjected to Lexogen polyA RNA Selection kit according to company recommendations. Enriched RNA has been used for sequencing library construction with CORALL Total RNA Seq Library Prep Kit according to the Lexogen manufacturer’s protocol. Zebrafish embryos were maintained according to standard protocol Westerfield et al. 2000 and staged according to standard morphological criteria Kimmel et al. 1995. strain:ABTL Tgddx4:ddx4 EGFP|tissue:whole embryo|condition:Mock|developmental stage:5 hpf GSM4407869 GSM4407869: Mock 5hpf Rep 3; Danio rerio; RNA Seq GSM4407869   1 25 embryos each for both StCsm and mock injected were pooled at xxx cell stage 5 hpf and 24 hpf from xxx biological replicates. Total RNA was extracted from the whole embryos with Tri Reagent Solution Invitrogen according to the manufacturer's protocol. RNA was further purified using Zymo RNA Clean and Concentrator 5 kit according to manufacturer's protocol. 1.5 ug of total RNA was subjected to Lexogen polyA RNA Selection kit according to company recommendations. Enriched RNA has been used for sequencing library construction with CORALL Total RNA Seq Library Prep Kit according to the Lexogen manufacturer's protocol. GEO Accession:GSM4407869 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP252498     Mock-5hpf-Rep-3_R1.fastq.gz Mock-5hpf-Rep-3_R2.fastq.gz fastq fastq 2842544510.0 21865727.0 GSM4407869 r1 0:71 1:59 A:799747787;C:578455085;G:650943235;T:809456844;N:3941559 71 59     799747787 578455085 650943235 809456844 3941559 SRX7899072 SRS6307327 SRA1054182 GEO Laboratory of Zebrafish Developmental Genomics, International Institute of Molecular and Cell Biology in Warsaw 2 0.60476 0.47987 0.07746 0.06523 0.76978 0.79064 0.53156 0.5417 71 59 B B biological fallback assumption illumina nextseq unknown cdna_unspecified lexogen bulk bulk bulk   Poland 2020-03-12 Blastula Embryo Whole Organism All anatomical structures
57242 57242 SRR11293509 SRX7899071 SRS6307331 SRP252498 PRJNA612134 Targeted RNA Knockdown by a Type III CRISPR Cas Complex in Zebrafish GSE146852 Transcriptome Analysis RNA interference RNAi is a powerful experimental tool for RNA knockdown but not all organisms are amenable. Here we provide a “proof of principle” demonstration that CRISPR endoribonuclease can be used for programmable mRNA transcript degradation in eukaryotes. Using zebrafish as the animal model and Csm complex as the CRISPR endoribonuclease we targeted EGFP transcript expressed from a variety of promoters. A drastic decrease of fluorescence was achieved in germ cells of the vasa:EGFP line. Weaker effects were also seen in fish lines that express EGFP zygotically. Knockdown was statistically significant in cmcl2:EGFP and fli1:EGFP zebrafish lines at 1 dpf dpf but reduced to background levels at 2 dpf. The nkx2.5:EGFP fish line was least susceptible to Csm mediated EGFP knockdown. We also tested Csm mediated knockdown on the endogenous tdgf1 oep transcript. At optimal Csm dose we observed a penetrance of the characteristic one eyed phenotype at greater than 50% penetrance and hence with similar efficiency to morpholino mediated knockdown. We conclude that Csm mediated knockdown is very efficient for maternal transcripts and can also be used for mixed maternal/early zygotic and early zygotic transcripts in some cases reaching comparable efficiency to morpholino based knockdown with no significant off target effects in our model. Overall design: Zebrafish embryos from the transgenic line Tgddx4:ddx4 EGFP on the background of ABTL were injected with the Csm complex targeted against the EGFP transcript or mock solution StCsm buffer without xxx proteins. The RNA was isolated from the pooled embryos at three time points: 128 cell stage 5 hpf and 24 hpf. Three replicates were obtained from each experimental condition and time point. Pair end sequencing was performed on the Illumina NextSeq platform.   pubmed:32833532   Mock 5hpf Rep 2 GSM4407868   source name:whole zebrafish embryo|strain:ABTL Tgddx4:ddx4 EGFP|tissue:whole embryo|condition:Mock|developmental stage:5 hpf Mock 5hpf Rep 2 Base calling was performed by Illumina RTA 1.18.61. Files were translated into demultiplexed FASTQ files by Illumina bcl2fastq2 2.16.10. The first 12 nucleotides of read 1 containing the UMI sequence were extracted and added to the sequence identifier using UMI tools v. 1.0.0. Low quality sequences and adapter content were trimmed with cutadapt v. 2.4. Reads were aligned to the GRCz11 reference genome supplemented with the EGFP sequence with STAR v. 2.7.2b using the following custom parameters: outFilterType BySJout outFilterMultimapNmax 20 alignSJoverhangMin 8 alignSJDBoverhangMin 1 outFilterMismatchNmax 999 alignIntronMin 20 alignIntronMax 1000000 alignMatesGapMax 1000000 outFilterMismatchNoverLmax 0.6. Alignments were de duplicated based on their Unique Molecular Identifier sequence using UMI tools v. 1.0.0. De duplicated alignments were counted per exon on the gene level with the htseq count script from the HTSeq v. 0.11.2 package using Ensembl GRCz11 version 97 genome annotation. Further analysis was performed in R v. 3.6.1 programming language. Differential expression was analyzed using DESeq2 v. 1.24.0 package. Genome build: GRCz11 Supplementary files format and content: HTSeq counts.tsv files were generated with htseq count script they contain counts for each gene for each sample in tab delimited format; normalized counts files were generated with DESeq2 they contain library size normalized read counts for each gene for every sample from each time point in tab delimited format; egfp.fa file contains the EGFP sequence in FASTA format; egfp.gtf file contains the custom EGFP annotation in GTF format; egfp crrna.fa file contains the crRNA sequence against the EGFP transcript used in the experiment in FASTA format. whole zebrafish embryo Zebrafish embryos from three different mating pairs of Tgddx4:ddx4 EGFP females with casper males were pooled during collection. D16A StCsmEGFP was diluted with water for injection 1:1 v/v up to a concentration of 0.7mg/ml. For mock injection Tris storage buffer 10mM Tris HCl pH8.5 300mM Nacl 1mM DTT 0.1mM EDTA 50% glycerol was also diluted in the similar manner. 1nl of either StCsm EGFP solution 0.7ng or buffer alone were injected into the yolk of embryos at the 1 cell stage. In total three independent injections were performed. 25 embryos for both StCsm and mock injected per replica set per time point were frozen in liquid nitrogen stored at xxx℃ and processed together. 25 embryos each for both StCsm and mock injected were pooled at xxx cell stage 5 hpf and 24 hpf from xxx biological replicates. Total RNA was extracted from the whole embryos with Tri Reagent Solution Invitrogen according to the manufacturer’s protocol. RNA was further purified using Zymo RNA Clean and Concentrator 5 kit according to manufacturer's protocol. 1.5 ug of total RNA was subjected to Lexogen polyA RNA Selection kit according to company recommendations. Enriched RNA has been used for sequencing library construction with CORALL Total RNA Seq Library Prep Kit according to the Lexogen manufacturer’s protocol. Zebrafish embryos were maintained according to standard protocol Westerfield et al. 2000 and staged according to standard morphological criteria Kimmel et al. 1995. strain:ABTL Tgddx4:ddx4 EGFP|tissue:whole embryo|condition:Mock|developmental stage:5 hpf GSM4407868 GSM4407868: Mock 5hpf Rep 2; Danio rerio; RNA Seq GSM4407868   1 25 embryos each for both StCsm and mock injected were pooled at xxx cell stage 5 hpf and 24 hpf from xxx biological replicates. Total RNA was extracted from the whole embryos with Tri Reagent Solution Invitrogen according to the manufacturer's protocol. RNA was further purified using Zymo RNA Clean and Concentrator 5 kit according to manufacturer's protocol. 1.5 ug of total RNA was subjected to Lexogen polyA RNA Selection kit according to company recommendations. Enriched RNA has been used for sequencing library construction with CORALL Total RNA Seq Library Prep Kit according to the Lexogen manufacturer's protocol. GEO Accession:GSM4407868 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP252498     Mock-5hpf-Rep-2_R2.fastq.gz Mock-5hpf-Rep-2_R1.fastq.gz fastq fastq 2568887880.0 19760676.0 GSM4407868 r1 0:71 1:59 A:734167827;C:503406924;G:571409098;T:756330561;N:3573470 71 59     734167827 503406924 571409098 756330561 3573470 SRX7899071 SRS6307331 SRA1054182 GEO Laboratory of Zebrafish Developmental Genomics, International Institute of Molecular and Cell Biology in Warsaw 2 0.65779 0.50537 0.10242 0.08466 0.76759 0.79492 0.50601 0.52295 71 59 B B biological fallback assumption illumina nextseq unknown cdna_unspecified lexogen bulk bulk bulk   Poland 2020-03-12 Blastula Embryo Whole Organism All anatomical structures
57243 57243 SRR11293508 SRX7899070 SRS6307326 SRP252498 PRJNA612134 Targeted RNA Knockdown by a Type III CRISPR Cas Complex in Zebrafish GSE146852 Transcriptome Analysis RNA interference RNAi is a powerful experimental tool for RNA knockdown but not all organisms are amenable. Here we provide a “proof of principle” demonstration that CRISPR endoribonuclease can be used for programmable mRNA transcript degradation in eukaryotes. Using zebrafish as the animal model and Csm complex as the CRISPR endoribonuclease we targeted EGFP transcript expressed from a variety of promoters. A drastic decrease of fluorescence was achieved in germ cells of the vasa:EGFP line. Weaker effects were also seen in fish lines that express EGFP zygotically. Knockdown was statistically significant in cmcl2:EGFP and fli1:EGFP zebrafish lines at 1 dpf dpf but reduced to background levels at 2 dpf. The nkx2.5:EGFP fish line was least susceptible to Csm mediated EGFP knockdown. We also tested Csm mediated knockdown on the endogenous tdgf1 oep transcript. At optimal Csm dose we observed a penetrance of the characteristic one eyed phenotype at greater than 50% penetrance and hence with similar efficiency to morpholino mediated knockdown. We conclude that Csm mediated knockdown is very efficient for maternal transcripts and can also be used for mixed maternal/early zygotic and early zygotic transcripts in some cases reaching comparable efficiency to morpholino based knockdown with no significant off target effects in our model. Overall design: Zebrafish embryos from the transgenic line Tgddx4:ddx4 EGFP on the background of ABTL were injected with the Csm complex targeted against the EGFP transcript or mock solution StCsm buffer without xxx proteins. The RNA was isolated from the pooled embryos at three time points: 128 cell stage 5 hpf and 24 hpf. Three replicates were obtained from each experimental condition and time point. Pair end sequencing was performed on the Illumina NextSeq platform.   pubmed:32833532   Mock 5hpf Rep 1 GSM4407867   source name:whole zebrafish embryo|strain:ABTL Tgddx4:ddx4 EGFP|tissue:whole embryo|condition:Mock|developmental stage:5 hpf Mock 5hpf Rep 1 Base calling was performed by Illumina RTA 1.18.61. Files were translated into demultiplexed FASTQ files by Illumina bcl2fastq2 2.16.10. The first 12 nucleotides of read 1 containing the UMI sequence were extracted and added to the sequence identifier using UMI tools v. 1.0.0. Low quality sequences and adapter content were trimmed with cutadapt v. 2.4. Reads were aligned to the GRCz11 reference genome supplemented with the EGFP sequence with STAR v. 2.7.2b using the following custom parameters: outFilterType BySJout outFilterMultimapNmax 20 alignSJoverhangMin 8 alignSJDBoverhangMin 1 outFilterMismatchNmax 999 alignIntronMin 20 alignIntronMax 1000000 alignMatesGapMax 1000000 outFilterMismatchNoverLmax 0.6. Alignments were de duplicated based on their Unique Molecular Identifier sequence using UMI tools v. 1.0.0. De duplicated alignments were counted per exon on the gene level with the htseq count script from the HTSeq v. 0.11.2 package using Ensembl GRCz11 version 97 genome annotation. Further analysis was performed in R v. 3.6.1 programming language. Differential expression was analyzed using DESeq2 v. 1.24.0 package. Genome build: GRCz11 Supplementary files format and content: HTSeq counts.tsv files were generated with htseq count script they contain counts for each gene for each sample in tab delimited format; normalized counts files were generated with DESeq2 they contain library size normalized read counts for each gene for every sample from each time point in tab delimited format; egfp.fa file contains the EGFP sequence in FASTA format; egfp.gtf file contains the custom EGFP annotation in GTF format; egfp crrna.fa file contains the crRNA sequence against the EGFP transcript used in the experiment in FASTA format. whole zebrafish embryo Zebrafish embryos from three different mating pairs of Tgddx4:ddx4 EGFP females with casper males were pooled during collection. D16A StCsmEGFP was diluted with water for injection 1:1 v/v up to a concentration of 0.7mg/ml. For mock injection Tris storage buffer 10mM Tris HCl pH8.5 300mM Nacl 1mM DTT 0.1mM EDTA 50% glycerol was also diluted in the similar manner. 1nl of either StCsm EGFP solution 0.7ng or buffer alone were injected into the yolk of embryos at the 1 cell stage. In total three independent injections were performed. 25 embryos for both StCsm and mock injected per replica set per time point were frozen in liquid nitrogen stored at xxx℃ and processed together. 25 embryos each for both StCsm and mock injected were pooled at xxx cell stage 5 hpf and 24 hpf from xxx biological replicates. Total RNA was extracted from the whole embryos with Tri Reagent Solution Invitrogen according to the manufacturer’s protocol. RNA was further purified using Zymo RNA Clean and Concentrator 5 kit according to manufacturer's protocol. 1.5 ug of total RNA was subjected to Lexogen polyA RNA Selection kit according to company recommendations. Enriched RNA has been used for sequencing library construction with CORALL Total RNA Seq Library Prep Kit according to the Lexogen manufacturer’s protocol. Zebrafish embryos were maintained according to standard protocol Westerfield et al. 2000 and staged according to standard morphological criteria Kimmel et al. 1995. strain:ABTL Tgddx4:ddx4 EGFP|tissue:whole embryo|condition:Mock|developmental stage:5 hpf GSM4407867 GSM4407867: Mock 5hpf Rep 1; Danio rerio; RNA Seq GSM4407867   1 25 embryos each for both StCsm and mock injected were pooled at xxx cell stage 5 hpf and 24 hpf from xxx biological replicates. Total RNA was extracted from the whole embryos with Tri Reagent Solution Invitrogen according to the manufacturer's protocol. RNA was further purified using Zymo RNA Clean and Concentrator 5 kit according to manufacturer's protocol. 1.5 ug of total RNA was subjected to Lexogen polyA RNA Selection kit according to company recommendations. Enriched RNA has been used for sequencing library construction with CORALL Total RNA Seq Library Prep Kit according to the Lexogen manufacturer's protocol. GEO Accession:GSM4407867 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP252498     Mock-5hpf-Rep-1_R1.fastq.gz Mock-5hpf-Rep-1_R2.fastq.gz fastq fastq 3252464540.0 25018958.0 GSM4407867 r1 0:71 1:59 A:898913139;C:680806953;G:759240096;T:908987584;N:4516768 71 59     898913139 680806953 759240096 908987584 4516768 SRX7899070 SRS6307326 SRA1054182 GEO Laboratory of Zebrafish Developmental Genomics, International Institute of Molecular and Cell Biology in Warsaw 2 0.60017 0.47729 0.08055 0.06599 0.78995 0.81126 0.57855 0.58317 71 59 B B biological fallback assumption illumina nextseq unknown cdna_unspecified lexogen bulk bulk bulk   Poland 2020-03-12 Blastula Embryo Whole Organism All anatomical structures
57247 57247 SRR11293504 SRX7899066 SRS6307322 SRP252498 PRJNA612134 Targeted RNA Knockdown by a Type III CRISPR Cas Complex in Zebrafish GSE146852 Transcriptome Analysis RNA interference RNAi is a powerful experimental tool for RNA knockdown but not all organisms are amenable. Here we provide a “proof of principle” demonstration that CRISPR endoribonuclease can be used for programmable mRNA transcript degradation in eukaryotes. Using zebrafish as the animal model and Csm complex as the CRISPR endoribonuclease we targeted EGFP transcript expressed from a variety of promoters. A drastic decrease of fluorescence was achieved in germ cells of the vasa:EGFP line. Weaker effects were also seen in fish lines that express EGFP zygotically. Knockdown was statistically significant in cmcl2:EGFP and fli1:EGFP zebrafish lines at 1 dpf dpf but reduced to background levels at 2 dpf. The nkx2.5:EGFP fish line was least susceptible to Csm mediated EGFP knockdown. We also tested Csm mediated knockdown on the endogenous tdgf1 oep transcript. At optimal Csm dose we observed a penetrance of the characteristic one eyed phenotype at greater than 50% penetrance and hence with similar efficiency to morpholino mediated knockdown. We conclude that Csm mediated knockdown is very efficient for maternal transcripts and can also be used for mixed maternal/early zygotic and early zygotic transcripts in some cases reaching comparable efficiency to morpholino based knockdown with no significant off target effects in our model. Overall design: Zebrafish embryos from the transgenic line Tgddx4:ddx4 EGFP on the background of ABTL were injected with the Csm complex targeted against the EGFP transcript or mock solution StCsm buffer without xxx proteins. The RNA was isolated from the pooled embryos at three time points: 128 cell stage 5 hpf and 24 hpf. Three replicates were obtained from each experimental condition and time point. Pair end sequencing was performed on the Illumina NextSeq platform.   pubmed:32833532   Mock 128 Rep 3 GSM4407863   source name:whole zebrafish embryo|strain:ABTL Tgddx4:ddx4 EGFP|tissue:whole embryo|condition:Mock|developmental stage:128 cells Mock 128 Rep 3 Base calling was performed by Illumina RTA 1.18.61. Files were translated into demultiplexed FASTQ files by Illumina bcl2fastq2 2.16.10. The first 12 nucleotides of read 1 containing the UMI sequence were extracted and added to the sequence identifier using UMI tools v. 1.0.0. Low quality sequences and adapter content were trimmed with cutadapt v. 2.4. Reads were aligned to the GRCz11 reference genome supplemented with the EGFP sequence with STAR v. 2.7.2b using the following custom parameters: outFilterType BySJout outFilterMultimapNmax 20 alignSJoverhangMin 8 alignSJDBoverhangMin 1 outFilterMismatchNmax 999 alignIntronMin 20 alignIntronMax 1000000 alignMatesGapMax 1000000 outFilterMismatchNoverLmax 0.6. Alignments were de duplicated based on their Unique Molecular Identifier sequence using UMI tools v. 1.0.0. De duplicated alignments were counted per exon on the gene level with the htseq count script from the HTSeq v. 0.11.2 package using Ensembl GRCz11 version 97 genome annotation. Further analysis was performed in R v. 3.6.1 programming language. Differential expression was analyzed using DESeq2 v. 1.24.0 package. Genome build: GRCz11 Supplementary files format and content: HTSeq counts.tsv files were generated with htseq count script they contain counts for each gene for each sample in tab delimited format; normalized counts files were generated with DESeq2 they contain library size normalized read counts for each gene for every sample from each time point in tab delimited format; egfp.fa file contains the EGFP sequence in FASTA format; egfp.gtf file contains the custom EGFP annotation in GTF format; egfp crrna.fa file contains the crRNA sequence against the EGFP transcript used in the experiment in FASTA format. whole zebrafish embryo Zebrafish embryos from three different mating pairs of Tgddx4:ddx4 EGFP females with casper males were pooled during collection. D16A StCsmEGFP was diluted with water for injection 1:1 v/v up to a concentration of 0.7mg/ml. For mock injection Tris storage buffer 10mM Tris HCl pH8.5 300mM Nacl 1mM DTT 0.1mM EDTA 50% glycerol was also diluted in the similar manner. 1nl of either StCsm EGFP solution 0.7ng or buffer alone were injected into the yolk of embryos at the 1 cell stage. In total three independent injections were performed. 25 embryos for both StCsm and mock injected per replica set per time point were frozen in liquid nitrogen stored at xxx℃ and processed together. 25 embryos each for both StCsm and mock injected were pooled at xxx cell stage 5 hpf and 24 hpf from xxx biological replicates. Total RNA was extracted from the whole embryos with Tri Reagent Solution Invitrogen according to the manufacturer’s protocol. RNA was further purified using Zymo RNA Clean and Concentrator 5 kit according to manufacturer's protocol. 1.5 ug of total RNA was subjected to Lexogen polyA RNA Selection kit according to company recommendations. Enriched RNA has been used for sequencing library construction with CORALL Total RNA Seq Library Prep Kit according to the Lexogen manufacturer’s protocol. Zebrafish embryos were maintained according to standard protocol Westerfield et al. 2000 and staged according to standard morphological criteria Kimmel et al. 1995. strain:ABTL Tgddx4:ddx4 EGFP|tissue:whole embryo|condition:Mock|developmental stage:128 cells GSM4407863 GSM4407863: Mock 128 Rep 3; Danio rerio; RNA Seq GSM4407863   1 25 embryos each for both StCsm and mock injected were pooled at xxx cell stage 5 hpf and 24 hpf from xxx biological replicates. Total RNA was extracted from the whole embryos with Tri Reagent Solution Invitrogen according to the manufacturer's protocol. RNA was further purified using Zymo RNA Clean and Concentrator 5 kit according to manufacturer's protocol. 1.5 ug of total RNA was subjected to Lexogen polyA RNA Selection kit according to company recommendations. Enriched RNA has been used for sequencing library construction with CORALL Total RNA Seq Library Prep Kit according to the Lexogen manufacturer's protocol. GEO Accession:GSM4407863 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP252498     Mock-128-Rep-3_R2.fastq.gz Mock-128-Rep-3_R1.fastq.gz fastq fastq 3261013600.0 25084720.0 GSM4407863 r1 0:71 1:59 A:900737410;C:688855174;G:770946078;T:895944416;N:4530522 71 59     900737410 688855174 770946078 895944416 4530522 SRX7899066 SRS6307322 SRA1054182 GEO Laboratory of Zebrafish Developmental Genomics, International Institute of Molecular and Cell Biology in Warsaw 2 0.57905 0.4659 0.03571 0.0302 0.77536 0.79022 0.5161 0.51323 71 59 B B biological fallback assumption illumina nextseq unknown cdna_unspecified lexogen bulk bulk bulk   Poland 2020-03-12 Blastula Embryo Whole Organism All anatomical structures
57248 57248 SRR11293503 SRX7899065 SRS6307321 SRP252498 PRJNA612134 Targeted RNA Knockdown by a Type III CRISPR Cas Complex in Zebrafish GSE146852 Transcriptome Analysis RNA interference RNAi is a powerful experimental tool for RNA knockdown but not all organisms are amenable. Here we provide a “proof of principle” demonstration that CRISPR endoribonuclease can be used for programmable mRNA transcript degradation in eukaryotes. Using zebrafish as the animal model and Csm complex as the CRISPR endoribonuclease we targeted EGFP transcript expressed from a variety of promoters. A drastic decrease of fluorescence was achieved in germ cells of the vasa:EGFP line. Weaker effects were also seen in fish lines that express EGFP zygotically. Knockdown was statistically significant in cmcl2:EGFP and fli1:EGFP zebrafish lines at 1 dpf dpf but reduced to background levels at 2 dpf. The nkx2.5:EGFP fish line was least susceptible to Csm mediated EGFP knockdown. We also tested Csm mediated knockdown on the endogenous tdgf1 oep transcript. At optimal Csm dose we observed a penetrance of the characteristic one eyed phenotype at greater than 50% penetrance and hence with similar efficiency to morpholino mediated knockdown. We conclude that Csm mediated knockdown is very efficient for maternal transcripts and can also be used for mixed maternal/early zygotic and early zygotic transcripts in some cases reaching comparable efficiency to morpholino based knockdown with no significant off target effects in our model. Overall design: Zebrafish embryos from the transgenic line Tgddx4:ddx4 EGFP on the background of ABTL were injected with the Csm complex targeted against the EGFP transcript or mock solution StCsm buffer without xxx proteins. The RNA was isolated from the pooled embryos at three time points: 128 cell stage 5 hpf and 24 hpf. Three replicates were obtained from each experimental condition and time point. Pair end sequencing was performed on the Illumina NextSeq platform.   pubmed:32833532   Mock 128 Rep 2 GSM4407862   source name:whole zebrafish embryo|strain:ABTL Tgddx4:ddx4 EGFP|tissue:whole embryo|condition:Mock|developmental stage:128 cells Mock 128 Rep 2 Base calling was performed by Illumina RTA 1.18.61. Files were translated into demultiplexed FASTQ files by Illumina bcl2fastq2 2.16.10. The first 12 nucleotides of read 1 containing the UMI sequence were extracted and added to the sequence identifier using UMI tools v. 1.0.0. Low quality sequences and adapter content were trimmed with cutadapt v. 2.4. Reads were aligned to the GRCz11 reference genome supplemented with the EGFP sequence with STAR v. 2.7.2b using the following custom parameters: outFilterType BySJout outFilterMultimapNmax 20 alignSJoverhangMin 8 alignSJDBoverhangMin 1 outFilterMismatchNmax 999 alignIntronMin 20 alignIntronMax 1000000 alignMatesGapMax 1000000 outFilterMismatchNoverLmax 0.6. Alignments were de duplicated based on their Unique Molecular Identifier sequence using UMI tools v. 1.0.0. De duplicated alignments were counted per exon on the gene level with the htseq count script from the HTSeq v. 0.11.2 package using Ensembl GRCz11 version 97 genome annotation. Further analysis was performed in R v. 3.6.1 programming language. Differential expression was analyzed using DESeq2 v. 1.24.0 package. Genome build: GRCz11 Supplementary files format and content: HTSeq counts.tsv files were generated with htseq count script they contain counts for each gene for each sample in tab delimited format; normalized counts files were generated with DESeq2 they contain library size normalized read counts for each gene for every sample from each time point in tab delimited format; egfp.fa file contains the EGFP sequence in FASTA format; egfp.gtf file contains the custom EGFP annotation in GTF format; egfp crrna.fa file contains the crRNA sequence against the EGFP transcript used in the experiment in FASTA format. whole zebrafish embryo Zebrafish embryos from three different mating pairs of Tgddx4:ddx4 EGFP females with casper males were pooled during collection. D16A StCsmEGFP was diluted with water for injection 1:1 v/v up to a concentration of 0.7mg/ml. For mock injection Tris storage buffer 10mM Tris HCl pH8.5 300mM Nacl 1mM DTT 0.1mM EDTA 50% glycerol was also diluted in the similar manner. 1nl of either StCsm EGFP solution 0.7ng or buffer alone were injected into the yolk of embryos at the 1 cell stage. In total three independent injections were performed. 25 embryos for both StCsm and mock injected per replica set per time point were frozen in liquid nitrogen stored at xxx℃ and processed together. 25 embryos each for both StCsm and mock injected were pooled at xxx cell stage 5 hpf and 24 hpf from xxx biological replicates. Total RNA was extracted from the whole embryos with Tri Reagent Solution Invitrogen according to the manufacturer’s protocol. RNA was further purified using Zymo RNA Clean and Concentrator 5 kit according to manufacturer's protocol. 1.5 ug of total RNA was subjected to Lexogen polyA RNA Selection kit according to company recommendations. Enriched RNA has been used for sequencing library construction with CORALL Total RNA Seq Library Prep Kit according to the Lexogen manufacturer’s protocol. Zebrafish embryos were maintained according to standard protocol Westerfield et al. 2000 and staged according to standard morphological criteria Kimmel et al. 1995. strain:ABTL Tgddx4:ddx4 EGFP|tissue:whole embryo|condition:Mock|developmental stage:128 cells GSM4407862 GSM4407862: Mock 128 Rep 2; Danio rerio; RNA Seq GSM4407862   1 25 embryos each for both StCsm and mock injected were pooled at xxx cell stage 5 hpf and 24 hpf from xxx biological replicates. Total RNA was extracted from the whole embryos with Tri Reagent Solution Invitrogen according to the manufacturer's protocol. RNA was further purified using Zymo RNA Clean and Concentrator 5 kit according to manufacturer's protocol. 1.5 ug of total RNA was subjected to Lexogen polyA RNA Selection kit according to company recommendations. Enriched RNA has been used for sequencing library construction with CORALL Total RNA Seq Library Prep Kit according to the Lexogen manufacturer's protocol. GEO Accession:GSM4407862 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP252498     Mock-128-Rep-2_R1.fastq.gz Mock-128-Rep-2_R2.fastq.gz fastq fastq 3077994140.0 23676878.0 GSM4407862 r1 0:71 1:59 A:848976357;C:649900868;G:727344485;T:847525958;N:4246472 71 59     848976357 649900868 727344485 847525958 4246472 SRX7899065 SRS6307321 SRA1054182 GEO Laboratory of Zebrafish Developmental Genomics, International Institute of Molecular and Cell Biology in Warsaw 2 0.56754 0.4574 0.03419 0.02915 0.77299 0.78934 0.5285 0.53073 71 59 B B biological fallback assumption illumina nextseq unknown cdna_unspecified lexogen bulk bulk bulk   Poland 2020-03-12 Blastula Embryo Whole Organism All anatomical structures
57249 57249 SRR11293502 SRX7899064 SRS6307320 SRP252498 PRJNA612134 Targeted RNA Knockdown by a Type III CRISPR Cas Complex in Zebrafish GSE146852 Transcriptome Analysis RNA interference RNAi is a powerful experimental tool for RNA knockdown but not all organisms are amenable. Here we provide a “proof of principle” demonstration that CRISPR endoribonuclease can be used for programmable mRNA transcript degradation in eukaryotes. Using zebrafish as the animal model and Csm complex as the CRISPR endoribonuclease we targeted EGFP transcript expressed from a variety of promoters. A drastic decrease of fluorescence was achieved in germ cells of the vasa:EGFP line. Weaker effects were also seen in fish lines that express EGFP zygotically. Knockdown was statistically significant in cmcl2:EGFP and fli1:EGFP zebrafish lines at 1 dpf dpf but reduced to background levels at 2 dpf. The nkx2.5:EGFP fish line was least susceptible to Csm mediated EGFP knockdown. We also tested Csm mediated knockdown on the endogenous tdgf1 oep transcript. At optimal Csm dose we observed a penetrance of the characteristic one eyed phenotype at greater than 50% penetrance and hence with similar efficiency to morpholino mediated knockdown. We conclude that Csm mediated knockdown is very efficient for maternal transcripts and can also be used for mixed maternal/early zygotic and early zygotic transcripts in some cases reaching comparable efficiency to morpholino based knockdown with no significant off target effects in our model. Overall design: Zebrafish embryos from the transgenic line Tgddx4:ddx4 EGFP on the background of ABTL were injected with the Csm complex targeted against the EGFP transcript or mock solution StCsm buffer without xxx proteins. The RNA was isolated from the pooled embryos at three time points: 128 cell stage 5 hpf and 24 hpf. Three replicates were obtained from each experimental condition and time point. Pair end sequencing was performed on the Illumina NextSeq platform.   pubmed:32833532   Mock 128 Rep 1 GSM4407861   source name:whole zebrafish embryo|strain:ABTL Tgddx4:ddx4 EGFP|tissue:whole embryo|condition:Mock|developmental stage:128 cells Mock 128 Rep 1 Base calling was performed by Illumina RTA 1.18.61. Files were translated into demultiplexed FASTQ files by Illumina bcl2fastq2 2.16.10. The first 12 nucleotides of read 1 containing the UMI sequence were extracted and added to the sequence identifier using UMI tools v. 1.0.0. Low quality sequences and adapter content were trimmed with cutadapt v. 2.4. Reads were aligned to the GRCz11 reference genome supplemented with the EGFP sequence with STAR v. 2.7.2b using the following custom parameters: outFilterType BySJout outFilterMultimapNmax 20 alignSJoverhangMin 8 alignSJDBoverhangMin 1 outFilterMismatchNmax 999 alignIntronMin 20 alignIntronMax 1000000 alignMatesGapMax 1000000 outFilterMismatchNoverLmax 0.6. Alignments were de duplicated based on their Unique Molecular Identifier sequence using UMI tools v. 1.0.0. De duplicated alignments were counted per exon on the gene level with the htseq count script from the HTSeq v. 0.11.2 package using Ensembl GRCz11 version 97 genome annotation. Further analysis was performed in R v. 3.6.1 programming language. Differential expression was analyzed using DESeq2 v. 1.24.0 package. Genome build: GRCz11 Supplementary files format and content: HTSeq counts.tsv files were generated with htseq count script they contain counts for each gene for each sample in tab delimited format; normalized counts files were generated with DESeq2 they contain library size normalized read counts for each gene for every sample from each time point in tab delimited format; egfp.fa file contains the EGFP sequence in FASTA format; egfp.gtf file contains the custom EGFP annotation in GTF format; egfp crrna.fa file contains the crRNA sequence against the EGFP transcript used in the experiment in FASTA format. whole zebrafish embryo Zebrafish embryos from three different mating pairs of Tgddx4:ddx4 EGFP females with casper males were pooled during collection. D16A StCsmEGFP was diluted with water for injection 1:1 v/v up to a concentration of 0.7mg/ml. For mock injection Tris storage buffer 10mM Tris HCl pH8.5 300mM Nacl 1mM DTT 0.1mM EDTA 50% glycerol was also diluted in the similar manner. 1nl of either StCsm EGFP solution 0.7ng or buffer alone were injected into the yolk of embryos at the 1 cell stage. In total three independent injections were performed. 25 embryos for both StCsm and mock injected per replica set per time point were frozen in liquid nitrogen stored at xxx℃ and processed together. 25 embryos each for both StCsm and mock injected were pooled at xxx cell stage 5 hpf and 24 hpf from xxx biological replicates. Total RNA was extracted from the whole embryos with Tri Reagent Solution Invitrogen according to the manufacturer’s protocol. RNA was further purified using Zymo RNA Clean and Concentrator 5 kit according to manufacturer's protocol. 1.5 ug of total RNA was subjected to Lexogen polyA RNA Selection kit according to company recommendations. Enriched RNA has been used for sequencing library construction with CORALL Total RNA Seq Library Prep Kit according to the Lexogen manufacturer’s protocol. Zebrafish embryos were maintained according to standard protocol Westerfield et al. 2000 and staged according to standard morphological criteria Kimmel et al. 1995. strain:ABTL Tgddx4:ddx4 EGFP|tissue:whole embryo|condition:Mock|developmental stage:128 cells GSM4407861 GSM4407861: Mock 128 Rep 1; Danio rerio; RNA Seq GSM4407861   1 25 embryos each for both StCsm and mock injected were pooled at xxx cell stage 5 hpf and 24 hpf from xxx biological replicates. Total RNA was extracted from the whole embryos with Tri Reagent Solution Invitrogen according to the manufacturer's protocol. RNA was further purified using Zymo RNA Clean and Concentrator 5 kit according to manufacturer's protocol. 1.5 ug of total RNA was subjected to Lexogen polyA RNA Selection kit according to company recommendations. Enriched RNA has been used for sequencing library construction with CORALL Total RNA Seq Library Prep Kit according to the Lexogen manufacturer's protocol. GEO Accession:GSM4407861 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA NextSeq 500   SRP252498     Mock-128-Rep-1_R1.fastq.gz Mock-128-Rep-1_R2.fastq.gz fastq fastq 2811933150.0 21630255.0 GSM4407861 r1 0:71 1:59 A:767935312;C:600889090;G:669149820;T:770070432;N:3888496 71 59     767935312 600889090 669149820 770070432 3888496 SRX7899064 SRS6307320 SRA1054182 GEO Laboratory of Zebrafish Developmental Genomics, International Institute of Molecular and Cell Biology in Warsaw 2 0.56804 0.46336 0.04038 0.03346 0.77638 0.79032 0.53645 0.52653 71 59 B B biological fallback assumption illumina nextseq unknown cdna_unspecified lexogen bulk bulk bulk   Poland 2020-03-12 Blastula Embryo Whole Organism All anatomical structures

Next page

Advanced export

JSON shape: default, array, newline-delimited

CSV options: 

CREATE TABLE run_metadata("run.accession" VARCHAR, "experiment.accession" VARCHAR, "sample.accession" VARCHAR, "study.accession" VARCHAR, bioproject VARCHAR, "study.title" VARCHAR, "study.alias" VARCHAR, "study.type" VARCHAR, "study.abstract" VARCHAR, "study.attributes" VARCHAR, "study.PMIDs" VARCHAR, "sample.description" VARCHAR, "sample.title" VARCHAR, "sample.alias" VARCHAR, "sample.centername" VARCHAR, "sample.attributes" VARCHAR, "GEOsample.title" VARCHAR, "GEOsample.dataprocessing" VARCHAR, "GEOsample.source" VARCHAR, "GEOsample.treatmentprotocol" VARCHAR, "GEOsample.extractprotocol" VARCHAR, "GEOsample.growthprotocol" VARCHAR, "GEOsample.characteristics" VARCHAR, "GEOsample.accession" VARCHAR, "experiment.title" VARCHAR, "experiment.alias" VARCHAR, "experiment.library_name" VARCHAR, "experiment.design_description" VARCHAR, "experiment.library_construction_protocol" VARCHAR, "experiment.attributes" VARCHAR, "experiment.library_strategy" VARCHAR, "experiment.library_source" VARCHAR, "experiment.library_selection" VARCHAR, "experiment.library_layout" VARCHAR, "experiment.platform" VARCHAR, "experiment.instrument_model" VARCHAR, "experiment.spot_descriptor" VARCHAR, "experiment.study_ref" VARCHAR, "run.title" VARCHAR, "run.attributes" VARCHAR, "run.filename" VARCHAR, "run.semantic_name" VARCHAR, "run.total_bases" DOUBLE, "run.total_spots" DOUBLE, "run.alias" VARCHAR, "run.read_lengths" VARCHAR, "run.base_counts" VARCHAR, "run.r1_length" BIGINT, "run.r2_length" BIGINT, "run.r3_length" BIGINT, "run.r4_length" BIGINT, "run.Acount" BIGINT, "run.Ccount" BIGINT, "run.Gcount" BIGINT, "run.Tcount" BIGINT, "run.Ncount" BIGINT, "run.experiment" VARCHAR, "run.pool_member" VARCHAR, "submission.accession" VARCHAR, "submission.srasource" VARCHAR, "submission.bioprojectsource" VARCHAR, "seqdetective.n_mates" BIGINT, "seqdetective.mapping_rate.mate1" DOUBLE, "seqdetective.mapping_rate.mate2" DOUBLE, "seqdetective.nofeature_rate.mate1" DOUBLE, "seqdetective.nofeature_rate.mate2" DOUBLE, "seqdetective.sparsity.mate1" DOUBLE, "seqdetective.sparsity.mate2" DOUBLE, "seqdetective.pos_strand_rate.mate1" DOUBLE, "seqdetective.pos_strand_rate.mate2" DOUBLE, "seqdetective.readlen.mate1" BIGINT, "seqdetective.readlen.mate2" BIGINT, "seqdetective.judgement.mate1" VARCHAR, "seqdetective.judgement.mate2" VARCHAR, "seqdetective.judgement.reason" VARCHAR, platform_family VARCHAR, instrument_generation VARCHAR, read_bias VARCHAR, selection_class VARCHAR, prep_kit VARCHAR, sc_or_bulk VARCHAR, tech_class VARCHAR, technology VARCHAR, tech_variant VARCHAR, "submission.bioprojectsource.country" VARCHAR, earliest_date DATE, devstage_curation VARCHAR, devstage_curation_coarse VARCHAR, tissue_curation VARCHAR, tissue_curation_coarse VARCHAR);;
CREATE INDEX idx_run_bioproject ON run_metadata(bioproject);;
CREATE INDEX idx_run_run_accession ON run_metadata("run.accession");;