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
25182,SRR25670729,SRX21396042,SRS18636200,SRP455680,PRJNA1006406,Control of polyA tail length and translation in vertebrate oocytes and early embryos,GSE241107,Other,During oocyte maturation and early embryonic development  polyA tail lengths strongly influence mRNA translation. However  how tail lengths are controlled at different developmental stages has been unclear. Here  we performed tail length and translational profiling of mRNA reporter libraries each with > 10 million three prime UTR sequence variants in frog oocytes and embryos  and fish embryos. These analyses revealed that the UUUUA motif specifies cytoplasmic polyadenylation and identified diverse context features that modulate the activity of this 5 mer. Additional sequence motifs drive stage specific deadenylation in embryos  and UUUUA and C rich motifs drive tail length independent translational repression in oocytes. A neural network model accurately predicts tail length change during oocyte maturation in frogs  mice  and humans. Analyses of human sequence variants showed that those predicted to disrupt tail length control have been under negative selection  implying that our insights into control of polyA tail length and translation have implications for human health and fertility. Overall design: Sythetic mRNA reporter libraries with random three prime UTR sequences under four different sequence contexts were injected into frog oocytes and embryos and fish embryos. PolyA tail lengths were measured to at different developmental stages to examine sequence motifs that caused tail length changes. At the same time  polyA tail lengths of endogenous mRNAs from frog oocytes and embryos  fish embryos  and mouse oocytes were measured to investiage how their tail lengths were controlled. Please note that sample titles have been updated on Jan 6  2024.,,,,Fish embryo mRNA germ ring PAL seq v4,GSM7716871,,source name:embryo|tissue:embryo|treatment:N1|geo loc name:missing|collection date:missing,Fish embryo mRNA germ ring PAL seq v4,For gene specific tail seq of mRNA reporters  data were processed with a custom script available at https://github.com/coffeebond/MPRA tail seq. For PAL seq v3 or v4  reads were trimmed with cutadapt v3.7 with the parameters “ m 15   quality base=64  q 20 20   match read wildcards  e 0.05  a NNNNATCTCGTATGCCGTCTTCTGCTTG  O 7”. The trimmed reads were mapped using STAR v2.7.1a to the reference database containing the genomic sequences of the organism from which the mRNAs were obtained  the genomic sequences of humans or fish  depending on which spike in RNAs were used  and the sequences of the polyA standards generated previously  with the parameters “  runThreadN 16   runMode alignReads   outFilterMultimapNmax 1   outReadsUnmapped Fastx   outFilterType BySJout   outSAMattributes All   outSAMtype BAM Unsorted SortedByCoordinate”. Uniquely mapped read were furhter processed with a custom script available at https://github.com/coffeebond/PAL seq. Assembly: Homo sapiens: GRCh38.p7 primary assembly. Mus muculus: GRCh38.p4  primary assembly. Xenopus laevis: v10.1 assembly. Danio rerio: GRCz11 assembly. Supplementary files format and content: For gene specific tail seq of mRNA reporters  tab delimited text files with columns indicating 1 sequence of the variable region; 2 median tail length; 3 number of reads; 4–7 tail lengths at quantile 10  25  75  and 90. Supplementary files format and content: For PAL seq v3 or v4  tab delimited files with columns indicating 1 gene ID or polyA site ID; 2 read cluster ID; 3 tail length Library strategy: PAL seq v4,embryo,,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer’s suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,tissue:embryo|treatment:N1,GSM7716871,GSM7716871: Fish embryo mRNA germ ring PAL seq v4; Danio rerio; OTHER,GSM7716871 r1,GSM7716871,1,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer's suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina HiSeq 2500,,SRP455680,,,Fish_embryo_mRNA_germ_ring_PAL_seq_v4_rep1_raw_read2.fastq.gz Fish_embryo_mRNA_germ_ring_PAL_seq_v4_rep1_raw_read1.fastq.gz,fastq fastq,2834842912.0,9234016.0,GSM7716871 r1,0:52 1:255,A:725811118;C:702410592;G:747583409;T:651018348;N:8019445,52,255,,,725811118,702410592,747583409,651018348,8019445,SRX21396042,SRS18636200,SRA1694849,Whitehead Institute,Whitehead Institute,2,0.00023,0.30494,8e-05,0.01793,0.99967,0.99971,0.5,1.0,52,255,T,B,mate1 technical by mapping diff,illumina,hiseq_era,unknown,poly_a,unknown,bulk,bulk,bulk,,United States,2023-08-17,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
25183,SRR25670730,SRX21396042,SRS18636200,SRP455680,PRJNA1006406,Control of polyA tail length and translation in vertebrate oocytes and early embryos,GSE241107,Other,During oocyte maturation and early embryonic development  polyA tail lengths strongly influence mRNA translation. However  how tail lengths are controlled at different developmental stages has been unclear. Here  we performed tail length and translational profiling of mRNA reporter libraries each with > 10 million three prime UTR sequence variants in frog oocytes and embryos  and fish embryos. These analyses revealed that the UUUUA motif specifies cytoplasmic polyadenylation and identified diverse context features that modulate the activity of this 5 mer. Additional sequence motifs drive stage specific deadenylation in embryos  and UUUUA and C rich motifs drive tail length independent translational repression in oocytes. A neural network model accurately predicts tail length change during oocyte maturation in frogs  mice  and humans. Analyses of human sequence variants showed that those predicted to disrupt tail length control have been under negative selection  implying that our insights into control of polyA tail length and translation have implications for human health and fertility. Overall design: Sythetic mRNA reporter libraries with random three prime UTR sequences under four different sequence contexts were injected into frog oocytes and embryos and fish embryos. PolyA tail lengths were measured to at different developmental stages to examine sequence motifs that caused tail length changes. At the same time  polyA tail lengths of endogenous mRNAs from frog oocytes and embryos  fish embryos  and mouse oocytes were measured to investiage how their tail lengths were controlled. Please note that sample titles have been updated on Jan 6  2024.,,,,Fish embryo mRNA germ ring PAL seq v4,GSM7716871,,source name:embryo|tissue:embryo|treatment:N1|geo loc name:missing|collection date:missing,Fish embryo mRNA germ ring PAL seq v4,For gene specific tail seq of mRNA reporters  data were processed with a custom script available at https://github.com/coffeebond/MPRA tail seq. For PAL seq v3 or v4  reads were trimmed with cutadapt v3.7 with the parameters “ m 15   quality base=64  q 20 20   match read wildcards  e 0.05  a NNNNATCTCGTATGCCGTCTTCTGCTTG  O 7”. The trimmed reads were mapped using STAR v2.7.1a to the reference database containing the genomic sequences of the organism from which the mRNAs were obtained  the genomic sequences of humans or fish  depending on which spike in RNAs were used  and the sequences of the polyA standards generated previously  with the parameters “  runThreadN 16   runMode alignReads   outFilterMultimapNmax 1   outReadsUnmapped Fastx   outFilterType BySJout   outSAMattributes All   outSAMtype BAM Unsorted SortedByCoordinate”. Uniquely mapped read were furhter processed with a custom script available at https://github.com/coffeebond/PAL seq. Assembly: Homo sapiens: GRCh38.p7 primary assembly. Mus muculus: GRCh38.p4  primary assembly. Xenopus laevis: v10.1 assembly. Danio rerio: GRCz11 assembly. Supplementary files format and content: For gene specific tail seq of mRNA reporters  tab delimited text files with columns indicating 1 sequence of the variable region; 2 median tail length; 3 number of reads; 4–7 tail lengths at quantile 10  25  75  and 90. Supplementary files format and content: For PAL seq v3 or v4  tab delimited files with columns indicating 1 gene ID or polyA site ID; 2 read cluster ID; 3 tail length Library strategy: PAL seq v4,embryo,,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer’s suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,tissue:embryo|treatment:N1,GSM7716871,GSM7716871: Fish embryo mRNA germ ring PAL seq v4; Danio rerio; OTHER,GSM7716871 r1,GSM7716871,1,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer's suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina HiSeq 2500,,SRP455680,,,Fish_embryo_mRNA_germ_ring_PAL_seq_v4_rep2_raw_read1.fastq.gz Fish_embryo_mRNA_germ_ring_PAL_seq_v4_rep2_raw_read2.fastq.gz,fastq fastq,3473033898.0,11312814.0,GSM7716871 r2,0:52 1:255,A:853178471;C:899976159;G:962100712;T:750811461;N:6967095,52,255,,,853178471,899976159,962100712,750811461,6967095,SRX21396042,SRS18636200,SRA1694849,Whitehead Institute,Whitehead Institute,2,0.00049,0.0,0.00012,0.0,0.99941,1.0,0.64864,,52,255,T,T,mates < 9% mapping rate,illumina,hiseq_era,unknown,poly_a,unknown,bulk,bulk,bulk,,United States,2023-08-17,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
25184,SRR25670731,SRX21396041,SRS18636199,SRP455680,PRJNA1006406,Control of polyA tail length and translation in vertebrate oocytes and early embryos,GSE241107,Other,During oocyte maturation and early embryonic development  polyA tail lengths strongly influence mRNA translation. However  how tail lengths are controlled at different developmental stages has been unclear. Here  we performed tail length and translational profiling of mRNA reporter libraries each with > 10 million three prime UTR sequence variants in frog oocytes and embryos  and fish embryos. These analyses revealed that the UUUUA motif specifies cytoplasmic polyadenylation and identified diverse context features that modulate the activity of this 5 mer. Additional sequence motifs drive stage specific deadenylation in embryos  and UUUUA and C rich motifs drive tail length independent translational repression in oocytes. A neural network model accurately predicts tail length change during oocyte maturation in frogs  mice  and humans. Analyses of human sequence variants showed that those predicted to disrupt tail length control have been under negative selection  implying that our insights into control of polyA tail length and translation have implications for human health and fertility. Overall design: Sythetic mRNA reporter libraries with random three prime UTR sequences under four different sequence contexts were injected into frog oocytes and embryos and fish embryos. PolyA tail lengths were measured to at different developmental stages to examine sequence motifs that caused tail length changes. At the same time  polyA tail lengths of endogenous mRNAs from frog oocytes and embryos  fish embryos  and mouse oocytes were measured to investiage how their tail lengths were controlled. Please note that sample titles have been updated on Jan 6  2024.,,,,Fish embryo mRNA zfs:0000015 PAL seq v4,GSM7716870,,source name:embryo|tissue:embryo|treatment:N1|geo loc name:missing|collection date:missing,Fish embryo mRNA zfs:0000015 PAL seq v4,For gene specific tail seq of mRNA reporters  data were processed with a custom script available at https://github.com/coffeebond/MPRA tail seq. For PAL seq v3 or v4  reads were trimmed with cutadapt v3.7 with the parameters “ m 15   quality base=64  q 20 20   match read wildcards  e 0.05  a NNNNATCTCGTATGCCGTCTTCTGCTTG  O 7”. The trimmed reads were mapped using STAR v2.7.1a to the reference database containing the genomic sequences of the organism from which the mRNAs were obtained  the genomic sequences of humans or fish  depending on which spike in RNAs were used  and the sequences of the polyA standards generated previously  with the parameters “  runThreadN 16   runMode alignReads   outFilterMultimapNmax 1   outReadsUnmapped Fastx   outFilterType BySJout   outSAMattributes All   outSAMtype BAM Unsorted SortedByCoordinate”. Uniquely mapped read were furhter processed with a custom script available at https://github.com/coffeebond/PAL seq. Assembly: Homo sapiens: GRCh38.p7 primary assembly. Mus muculus: GRCh38.p4  primary assembly. Xenopus laevis: v10.1 assembly. Danio rerio: GRCz11 assembly. Supplementary files format and content: For gene specific tail seq of mRNA reporters  tab delimited text files with columns indicating 1 sequence of the variable region; 2 median tail length; 3 number of reads; 4–7 tail lengths at quantile 10  25  75  and 90. Supplementary files format and content: For PAL seq v3 or v4  tab delimited files with columns indicating 1 gene ID or polyA site ID; 2 read cluster ID; 3 tail length Library strategy: PAL seq v4,embryo,,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer’s suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,tissue:embryo|treatment:N1,GSM7716870,GSM7716870: Fish embryo mRNA zfs:0000015 PAL seq v4; Danio rerio; OTHER,GSM7716870 r1,GSM7716870,1,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer's suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina HiSeq 2500,,SRP455680,,,Fish_embryo_mRNA_30percent_epiboly_PAL_seq_v4_rep1_raw_read1.fastq.gz Fish_embryo_mRNA_30percent_epiboly_PAL_seq_v4_rep1_raw_read2.fastq.gz,fastq fastq,2695203962.0,8779166.0,GSM7716870 r1,0:52 1:255,A:684535386;C:669852151;G:719008886;T:614209811;N:7597728,52,255,,,684535386,669852151,719008886,614209811,7597728,SRX21396041,SRS18636199,SRA1694849,Whitehead Institute,Whitehead Institute,2,0.00117,0.35837,0.00017,0.00682,0.99859,0.99963,0.69473,1.0,52,255,T,B,mate1 technical by mapping diff,illumina,hiseq_era,unknown,poly_a,unknown,bulk,bulk,bulk,,United States,2023-08-17,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
25185,SRR25670732,SRX21396041,SRS18636199,SRP455680,PRJNA1006406,Control of polyA tail length and translation in vertebrate oocytes and early embryos,GSE241107,Other,During oocyte maturation and early embryonic development  polyA tail lengths strongly influence mRNA translation. However  how tail lengths are controlled at different developmental stages has been unclear. Here  we performed tail length and translational profiling of mRNA reporter libraries each with > 10 million three prime UTR sequence variants in frog oocytes and embryos  and fish embryos. These analyses revealed that the UUUUA motif specifies cytoplasmic polyadenylation and identified diverse context features that modulate the activity of this 5 mer. Additional sequence motifs drive stage specific deadenylation in embryos  and UUUUA and C rich motifs drive tail length independent translational repression in oocytes. A neural network model accurately predicts tail length change during oocyte maturation in frogs  mice  and humans. Analyses of human sequence variants showed that those predicted to disrupt tail length control have been under negative selection  implying that our insights into control of polyA tail length and translation have implications for human health and fertility. Overall design: Sythetic mRNA reporter libraries with random three prime UTR sequences under four different sequence contexts were injected into frog oocytes and embryos and fish embryos. PolyA tail lengths were measured to at different developmental stages to examine sequence motifs that caused tail length changes. At the same time  polyA tail lengths of endogenous mRNAs from frog oocytes and embryos  fish embryos  and mouse oocytes were measured to investiage how their tail lengths were controlled. Please note that sample titles have been updated on Jan 6  2024.,,,,Fish embryo mRNA zfs:0000015 PAL seq v4,GSM7716870,,source name:embryo|tissue:embryo|treatment:N1|geo loc name:missing|collection date:missing,Fish embryo mRNA zfs:0000015 PAL seq v4,For gene specific tail seq of mRNA reporters  data were processed with a custom script available at https://github.com/coffeebond/MPRA tail seq. For PAL seq v3 or v4  reads were trimmed with cutadapt v3.7 with the parameters “ m 15   quality base=64  q 20 20   match read wildcards  e 0.05  a NNNNATCTCGTATGCCGTCTTCTGCTTG  O 7”. The trimmed reads were mapped using STAR v2.7.1a to the reference database containing the genomic sequences of the organism from which the mRNAs were obtained  the genomic sequences of humans or fish  depending on which spike in RNAs were used  and the sequences of the polyA standards generated previously  with the parameters “  runThreadN 16   runMode alignReads   outFilterMultimapNmax 1   outReadsUnmapped Fastx   outFilterType BySJout   outSAMattributes All   outSAMtype BAM Unsorted SortedByCoordinate”. Uniquely mapped read were furhter processed with a custom script available at https://github.com/coffeebond/PAL seq. Assembly: Homo sapiens: GRCh38.p7 primary assembly. Mus muculus: GRCh38.p4  primary assembly. Xenopus laevis: v10.1 assembly. Danio rerio: GRCz11 assembly. Supplementary files format and content: For gene specific tail seq of mRNA reporters  tab delimited text files with columns indicating 1 sequence of the variable region; 2 median tail length; 3 number of reads; 4–7 tail lengths at quantile 10  25  75  and 90. Supplementary files format and content: For PAL seq v3 or v4  tab delimited files with columns indicating 1 gene ID or polyA site ID; 2 read cluster ID; 3 tail length Library strategy: PAL seq v4,embryo,,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer’s suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,tissue:embryo|treatment:N1,GSM7716870,GSM7716870: Fish embryo mRNA zfs:0000015 PAL seq v4; Danio rerio; OTHER,GSM7716870 r1,GSM7716870,1,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer's suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina HiSeq 2500,,SRP455680,,,Fish_embryo_mRNA_30percent_epiboly_PAL_seq_v4_rep2_raw_read2.fastq.gz Fish_embryo_mRNA_30percent_epiboly_PAL_seq_v4_rep2_raw_read1.fastq.gz,fastq fastq,3476338446.0,11323578.0,GSM7716870 r2,0:52 1:255,A:841832234;C:898325199;G:968774318;T:760398070;N:7008625,52,255,,,841832234,898325199,968774318,760398070,7008625,SRX21396041,SRS18636199,SRA1694849,Whitehead Institute,Whitehead Institute,2,0.00245,0.0,0.00047,0.0,0.99803,1.0,0.58536,,52,255,T,T,mates < 9% mapping rate,illumina,hiseq_era,unknown,poly_a,unknown,bulk,bulk,bulk,,United States,2023-08-17,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
25186,SRR25670733,SRX21396040,SRS18636198,SRP455680,PRJNA1006406,Control of polyA tail length and translation in vertebrate oocytes and early embryos,GSE241107,Other,During oocyte maturation and early embryonic development  polyA tail lengths strongly influence mRNA translation. However  how tail lengths are controlled at different developmental stages has been unclear. Here  we performed tail length and translational profiling of mRNA reporter libraries each with > 10 million three prime UTR sequence variants in frog oocytes and embryos  and fish embryos. These analyses revealed that the UUUUA motif specifies cytoplasmic polyadenylation and identified diverse context features that modulate the activity of this 5 mer. Additional sequence motifs drive stage specific deadenylation in embryos  and UUUUA and C rich motifs drive tail length independent translational repression in oocytes. A neural network model accurately predicts tail length change during oocyte maturation in frogs  mice  and humans. Analyses of human sequence variants showed that those predicted to disrupt tail length control have been under negative selection  implying that our insights into control of polyA tail length and translation have implications for human health and fertility. Overall design: Sythetic mRNA reporter libraries with random three prime UTR sequences under four different sequence contexts were injected into frog oocytes and embryos and fish embryos. PolyA tail lengths were measured to at different developmental stages to examine sequence motifs that caused tail length changes. At the same time  polyA tail lengths of endogenous mRNAs from frog oocytes and embryos  fish embryos  and mouse oocytes were measured to investiage how their tail lengths were controlled. Please note that sample titles have been updated on Jan 6  2024.,,,,Fish embryo mRNA sphere PAL seq v4,GSM7716869,,source name:embryo|tissue:embryo|treatment:N1|geo loc name:missing|collection date:missing,Fish embryo mRNA sphere PAL seq v4,For gene specific tail seq of mRNA reporters  data were processed with a custom script available at https://github.com/coffeebond/MPRA tail seq. For PAL seq v3 or v4  reads were trimmed with cutadapt v3.7 with the parameters “ m 15   quality base=64  q 20 20   match read wildcards  e 0.05  a NNNNATCTCGTATGCCGTCTTCTGCTTG  O 7”. The trimmed reads were mapped using STAR v2.7.1a to the reference database containing the genomic sequences of the organism from which the mRNAs were obtained  the genomic sequences of humans or fish  depending on which spike in RNAs were used  and the sequences of the polyA standards generated previously  with the parameters “  runThreadN 16   runMode alignReads   outFilterMultimapNmax 1   outReadsUnmapped Fastx   outFilterType BySJout   outSAMattributes All   outSAMtype BAM Unsorted SortedByCoordinate”. Uniquely mapped read were furhter processed with a custom script available at https://github.com/coffeebond/PAL seq. Assembly: Homo sapiens: GRCh38.p7 primary assembly. Mus muculus: GRCh38.p4  primary assembly. Xenopus laevis: v10.1 assembly. Danio rerio: GRCz11 assembly. Supplementary files format and content: For gene specific tail seq of mRNA reporters  tab delimited text files with columns indicating 1 sequence of the variable region; 2 median tail length; 3 number of reads; 4–7 tail lengths at quantile 10  25  75  and 90. Supplementary files format and content: For PAL seq v3 or v4  tab delimited files with columns indicating 1 gene ID or polyA site ID; 2 read cluster ID; 3 tail length Library strategy: PAL seq v4,embryo,,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer’s suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,tissue:embryo|treatment:N1,GSM7716869,GSM7716869: Fish embryo mRNA sphere PAL seq v4; Danio rerio; OTHER,GSM7716869 r1,GSM7716869,1,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer's suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina HiSeq 2500,,SRP455680,,,Fish_embryo_mRNA_sphere_PAL_seq_v4_rep1_raw_read1.fastq.gz Fish_embryo_mRNA_sphere_PAL_seq_v4_rep1_raw_read2.fastq.gz,fastq fastq,3211687254.0,10461522.0,GSM7716869 r1,0:52 1:255,A:813560400;C:775326762;G:854563477;T:759161458;N:9075157,52,255,,,813560400,775326762,854563477,759161458,9075157,SRX21396040,SRS18636198,SRA1694849,Whitehead Institute,Whitehead Institute,2,0.00088,0.43191,0.0004,0.01556,0.99916,0.99961,0.55769,1.0,52,255,T,B,mate1 technical by mapping diff,illumina,hiseq_era,unknown,poly_a,unknown,bulk,bulk,bulk,,United States,2023-08-17,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
25187,SRR25670734,SRX21396040,SRS18636198,SRP455680,PRJNA1006406,Control of polyA tail length and translation in vertebrate oocytes and early embryos,GSE241107,Other,During oocyte maturation and early embryonic development  polyA tail lengths strongly influence mRNA translation. However  how tail lengths are controlled at different developmental stages has been unclear. Here  we performed tail length and translational profiling of mRNA reporter libraries each with > 10 million three prime UTR sequence variants in frog oocytes and embryos  and fish embryos. These analyses revealed that the UUUUA motif specifies cytoplasmic polyadenylation and identified diverse context features that modulate the activity of this 5 mer. Additional sequence motifs drive stage specific deadenylation in embryos  and UUUUA and C rich motifs drive tail length independent translational repression in oocytes. A neural network model accurately predicts tail length change during oocyte maturation in frogs  mice  and humans. Analyses of human sequence variants showed that those predicted to disrupt tail length control have been under negative selection  implying that our insights into control of polyA tail length and translation have implications for human health and fertility. Overall design: Sythetic mRNA reporter libraries with random three prime UTR sequences under four different sequence contexts were injected into frog oocytes and embryos and fish embryos. PolyA tail lengths were measured to at different developmental stages to examine sequence motifs that caused tail length changes. At the same time  polyA tail lengths of endogenous mRNAs from frog oocytes and embryos  fish embryos  and mouse oocytes were measured to investiage how their tail lengths were controlled. Please note that sample titles have been updated on Jan 6  2024.,,,,Fish embryo mRNA sphere PAL seq v4,GSM7716869,,source name:embryo|tissue:embryo|treatment:N1|geo loc name:missing|collection date:missing,Fish embryo mRNA sphere PAL seq v4,For gene specific tail seq of mRNA reporters  data were processed with a custom script available at https://github.com/coffeebond/MPRA tail seq. For PAL seq v3 or v4  reads were trimmed with cutadapt v3.7 with the parameters “ m 15   quality base=64  q 20 20   match read wildcards  e 0.05  a NNNNATCTCGTATGCCGTCTTCTGCTTG  O 7”. The trimmed reads were mapped using STAR v2.7.1a to the reference database containing the genomic sequences of the organism from which the mRNAs were obtained  the genomic sequences of humans or fish  depending on which spike in RNAs were used  and the sequences of the polyA standards generated previously  with the parameters “  runThreadN 16   runMode alignReads   outFilterMultimapNmax 1   outReadsUnmapped Fastx   outFilterType BySJout   outSAMattributes All   outSAMtype BAM Unsorted SortedByCoordinate”. Uniquely mapped read were furhter processed with a custom script available at https://github.com/coffeebond/PAL seq. Assembly: Homo sapiens: GRCh38.p7 primary assembly. Mus muculus: GRCh38.p4  primary assembly. Xenopus laevis: v10.1 assembly. Danio rerio: GRCz11 assembly. Supplementary files format and content: For gene specific tail seq of mRNA reporters  tab delimited text files with columns indicating 1 sequence of the variable region; 2 median tail length; 3 number of reads; 4–7 tail lengths at quantile 10  25  75  and 90. Supplementary files format and content: For PAL seq v3 or v4  tab delimited files with columns indicating 1 gene ID or polyA site ID; 2 read cluster ID; 3 tail length Library strategy: PAL seq v4,embryo,,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer’s suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,tissue:embryo|treatment:N1,GSM7716869,GSM7716869: Fish embryo mRNA sphere PAL seq v4; Danio rerio; OTHER,GSM7716869 r1,GSM7716869,1,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer's suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina HiSeq 2500,,SRP455680,,,Fish_embryo_mRNA_sphere_PAL_seq_v4_rep2_raw_read1.fastq.gz Fish_embryo_mRNA_sphere_PAL_seq_v4_rep2_raw_read2.fastq.gz,fastq fastq,3179916131.0,10358033.0,GSM7716869 r2,0:52 1:255,A:771755667;C:804256300;G:883166175;T:714242073;N:6495916,52,255,,,771755667,804256300,883166175,714242073,6495916,SRX21396040,SRS18636198,SRA1694849,Whitehead Institute,Whitehead Institute,2,0.00186,0.0,0.00088,0.0,0.99862,1.0,0.64705,,52,255,T,T,mates < 9% mapping rate,illumina,hiseq_era,unknown,poly_a,unknown,bulk,bulk,bulk,,United States,2023-08-17,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
25188,SRR25670735,SRX21396039,SRS18636197,SRP455680,PRJNA1006406,Control of polyA tail length and translation in vertebrate oocytes and early embryos,GSE241107,Other,During oocyte maturation and early embryonic development  polyA tail lengths strongly influence mRNA translation. However  how tail lengths are controlled at different developmental stages has been unclear. Here  we performed tail length and translational profiling of mRNA reporter libraries each with > 10 million three prime UTR sequence variants in frog oocytes and embryos  and fish embryos. These analyses revealed that the UUUUA motif specifies cytoplasmic polyadenylation and identified diverse context features that modulate the activity of this 5 mer. Additional sequence motifs drive stage specific deadenylation in embryos  and UUUUA and C rich motifs drive tail length independent translational repression in oocytes. A neural network model accurately predicts tail length change during oocyte maturation in frogs  mice  and humans. Analyses of human sequence variants showed that those predicted to disrupt tail length control have been under negative selection  implying that our insights into control of polyA tail length and translation have implications for human health and fertility. Overall design: Sythetic mRNA reporter libraries with random three prime UTR sequences under four different sequence contexts were injected into frog oocytes and embryos and fish embryos. PolyA tail lengths were measured to at different developmental stages to examine sequence motifs that caused tail length changes. At the same time  polyA tail lengths of endogenous mRNAs from frog oocytes and embryos  fish embryos  and mouse oocytes were measured to investiage how their tail lengths were controlled. Please note that sample titles have been updated on Jan 6  2024.,,,,Fish embryo mRNA 1024cell PAL seq v4,GSM7716868,,source name:embryo|tissue:embryo|treatment:N1|geo loc name:missing|collection date:missing,Fish embryo mRNA 1024cell PAL seq v4,For gene specific tail seq of mRNA reporters  data were processed with a custom script available at https://github.com/coffeebond/MPRA tail seq. For PAL seq v3 or v4  reads were trimmed with cutadapt v3.7 with the parameters “ m 15   quality base=64  q 20 20   match read wildcards  e 0.05  a NNNNATCTCGTATGCCGTCTTCTGCTTG  O 7”. The trimmed reads were mapped using STAR v2.7.1a to the reference database containing the genomic sequences of the organism from which the mRNAs were obtained  the genomic sequences of humans or fish  depending on which spike in RNAs were used  and the sequences of the polyA standards generated previously  with the parameters “  runThreadN 16   runMode alignReads   outFilterMultimapNmax 1   outReadsUnmapped Fastx   outFilterType BySJout   outSAMattributes All   outSAMtype BAM Unsorted SortedByCoordinate”. Uniquely mapped read were furhter processed with a custom script available at https://github.com/coffeebond/PAL seq. Assembly: Homo sapiens: GRCh38.p7 primary assembly. Mus muculus: GRCh38.p4  primary assembly. Xenopus laevis: v10.1 assembly. Danio rerio: GRCz11 assembly. Supplementary files format and content: For gene specific tail seq of mRNA reporters  tab delimited text files with columns indicating 1 sequence of the variable region; 2 median tail length; 3 number of reads; 4–7 tail lengths at quantile 10  25  75  and 90. Supplementary files format and content: For PAL seq v3 or v4  tab delimited files with columns indicating 1 gene ID or polyA site ID; 2 read cluster ID; 3 tail length Library strategy: PAL seq v4,embryo,,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer’s suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,tissue:embryo|treatment:N1,GSM7716868,GSM7716868: Fish embryo mRNA 1024cell PAL seq v4; Danio rerio; OTHER,GSM7716868 r1,GSM7716868,1,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer's suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina HiSeq 2500,,SRP455680,,,Fish_embryo_mRNA_1024cell_PAL_seq_v4_rep1_raw_read1.fastq.gz Fish_embryo_mRNA_1024cell_PAL_seq_v4_rep1_raw_read2.fastq.gz,fastq fastq,2191900794.0,7139742.0,GSM7716868 r1,0:52 1:255,A:571954354;C:551162617;G:572445400;T:490238669;N:6099754,52,255,,,571954354,551162617,572445400,490238669,6099754,SRX21396039,SRS18636197,SRA1694849,Whitehead Institute,Whitehead Institute,2,0.0011,0.43387,6e-05,0.01058,0.99864,0.99971,0.74576,1.0,52,255,T,B,mate1 technical by mapping diff,illumina,hiseq_era,unknown,poly_a,unknown,bulk,bulk,bulk,,United States,2023-08-17,Zygote,Embryo,Embryo Imprecise,All anatomical structures
25189,SRR25670736,SRX21396039,SRS18636197,SRP455680,PRJNA1006406,Control of polyA tail length and translation in vertebrate oocytes and early embryos,GSE241107,Other,During oocyte maturation and early embryonic development  polyA tail lengths strongly influence mRNA translation. However  how tail lengths are controlled at different developmental stages has been unclear. Here  we performed tail length and translational profiling of mRNA reporter libraries each with > 10 million three prime UTR sequence variants in frog oocytes and embryos  and fish embryos. These analyses revealed that the UUUUA motif specifies cytoplasmic polyadenylation and identified diverse context features that modulate the activity of this 5 mer. Additional sequence motifs drive stage specific deadenylation in embryos  and UUUUA and C rich motifs drive tail length independent translational repression in oocytes. A neural network model accurately predicts tail length change during oocyte maturation in frogs  mice  and humans. Analyses of human sequence variants showed that those predicted to disrupt tail length control have been under negative selection  implying that our insights into control of polyA tail length and translation have implications for human health and fertility. Overall design: Sythetic mRNA reporter libraries with random three prime UTR sequences under four different sequence contexts were injected into frog oocytes and embryos and fish embryos. PolyA tail lengths were measured to at different developmental stages to examine sequence motifs that caused tail length changes. At the same time  polyA tail lengths of endogenous mRNAs from frog oocytes and embryos  fish embryos  and mouse oocytes were measured to investiage how their tail lengths were controlled. Please note that sample titles have been updated on Jan 6  2024.,,,,Fish embryo mRNA 1024cell PAL seq v4,GSM7716868,,source name:embryo|tissue:embryo|treatment:N1|geo loc name:missing|collection date:missing,Fish embryo mRNA 1024cell PAL seq v4,For gene specific tail seq of mRNA reporters  data were processed with a custom script available at https://github.com/coffeebond/MPRA tail seq. For PAL seq v3 or v4  reads were trimmed with cutadapt v3.7 with the parameters “ m 15   quality base=64  q 20 20   match read wildcards  e 0.05  a NNNNATCTCGTATGCCGTCTTCTGCTTG  O 7”. The trimmed reads were mapped using STAR v2.7.1a to the reference database containing the genomic sequences of the organism from which the mRNAs were obtained  the genomic sequences of humans or fish  depending on which spike in RNAs were used  and the sequences of the polyA standards generated previously  with the parameters “  runThreadN 16   runMode alignReads   outFilterMultimapNmax 1   outReadsUnmapped Fastx   outFilterType BySJout   outSAMattributes All   outSAMtype BAM Unsorted SortedByCoordinate”. Uniquely mapped read were furhter processed with a custom script available at https://github.com/coffeebond/PAL seq. Assembly: Homo sapiens: GRCh38.p7 primary assembly. Mus muculus: GRCh38.p4  primary assembly. Xenopus laevis: v10.1 assembly. Danio rerio: GRCz11 assembly. Supplementary files format and content: For gene specific tail seq of mRNA reporters  tab delimited text files with columns indicating 1 sequence of the variable region; 2 median tail length; 3 number of reads; 4–7 tail lengths at quantile 10  25  75  and 90. Supplementary files format and content: For PAL seq v3 or v4  tab delimited files with columns indicating 1 gene ID or polyA site ID; 2 read cluster ID; 3 tail length Library strategy: PAL seq v4,embryo,,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer’s suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,tissue:embryo|treatment:N1,GSM7716868,GSM7716868: Fish embryo mRNA 1024cell PAL seq v4; Danio rerio; OTHER,GSM7716868 r1,GSM7716868,1,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer's suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina HiSeq 2500,,SRP455680,,,Fish_embryo_mRNA_1024cell_PAL_seq_v4_rep2_raw_read1.fastq.gz Fish_embryo_mRNA_1024cell_PAL_seq_v4_rep2_raw_read2.fastq.gz,fastq fastq,3840723193.0,12510499.0,GSM7716868 r2,0:52 1:255,A:985340216;C:991311706;G:1034487140;T:821820974;N:7763157,52,255,,,985340216,991311706,1034487140,821820974,7763157,SRX21396039,SRS18636197,SRA1694849,Whitehead Institute,Whitehead Institute,2,0.0021,0.0,0.00026,0.0,0.99859,1.0,0.71022,,52,255,T,T,mates < 9% mapping rate,illumina,hiseq_era,unknown,poly_a,unknown,bulk,bulk,bulk,,United States,2023-08-17,Zygote,Embryo,Embryo Imprecise,All anatomical structures
25190,SRR25670737,SRX21396038,SRS18636196,SRP455680,PRJNA1006406,Control of polyA tail length and translation in vertebrate oocytes and early embryos,GSE241107,Other,During oocyte maturation and early embryonic development  polyA tail lengths strongly influence mRNA translation. However  how tail lengths are controlled at different developmental stages has been unclear. Here  we performed tail length and translational profiling of mRNA reporter libraries each with > 10 million three prime UTR sequence variants in frog oocytes and embryos  and fish embryos. These analyses revealed that the UUUUA motif specifies cytoplasmic polyadenylation and identified diverse context features that modulate the activity of this 5 mer. Additional sequence motifs drive stage specific deadenylation in embryos  and UUUUA and C rich motifs drive tail length independent translational repression in oocytes. A neural network model accurately predicts tail length change during oocyte maturation in frogs  mice  and humans. Analyses of human sequence variants showed that those predicted to disrupt tail length control have been under negative selection  implying that our insights into control of polyA tail length and translation have implications for human health and fertility. Overall design: Sythetic mRNA reporter libraries with random three prime UTR sequences under four different sequence contexts were injected into frog oocytes and embryos and fish embryos. PolyA tail lengths were measured to at different developmental stages to examine sequence motifs that caused tail length changes. At the same time  polyA tail lengths of endogenous mRNAs from frog oocytes and embryos  fish embryos  and mouse oocytes were measured to investiage how their tail lengths were controlled. Please note that sample titles have been updated on Jan 6  2024.,,,,Fish embryo mRNA 128cell PAL seq v4,GSM7716867,,source name:embryo|tissue:embryo|treatment:N1|geo loc name:missing|collection date:missing,Fish embryo mRNA 128cell PAL seq v4,For gene specific tail seq of mRNA reporters  data were processed with a custom script available at https://github.com/coffeebond/MPRA tail seq. For PAL seq v3 or v4  reads were trimmed with cutadapt v3.7 with the parameters “ m 15   quality base=64  q 20 20   match read wildcards  e 0.05  a NNNNATCTCGTATGCCGTCTTCTGCTTG  O 7”. The trimmed reads were mapped using STAR v2.7.1a to the reference database containing the genomic sequences of the organism from which the mRNAs were obtained  the genomic sequences of humans or fish  depending on which spike in RNAs were used  and the sequences of the polyA standards generated previously  with the parameters “  runThreadN 16   runMode alignReads   outFilterMultimapNmax 1   outReadsUnmapped Fastx   outFilterType BySJout   outSAMattributes All   outSAMtype BAM Unsorted SortedByCoordinate”. Uniquely mapped read were furhter processed with a custom script available at https://github.com/coffeebond/PAL seq. Assembly: Homo sapiens: GRCh38.p7 primary assembly. Mus muculus: GRCh38.p4  primary assembly. Xenopus laevis: v10.1 assembly. Danio rerio: GRCz11 assembly. Supplementary files format and content: For gene specific tail seq of mRNA reporters  tab delimited text files with columns indicating 1 sequence of the variable region; 2 median tail length; 3 number of reads; 4–7 tail lengths at quantile 10  25  75  and 90. Supplementary files format and content: For PAL seq v3 or v4  tab delimited files with columns indicating 1 gene ID or polyA site ID; 2 read cluster ID; 3 tail length Library strategy: PAL seq v4,embryo,,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer’s suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,tissue:embryo|treatment:N1,GSM7716867,GSM7716867: Fish embryo mRNA 128cell PAL seq v4; Danio rerio; OTHER,GSM7716867 r1,GSM7716867,1,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer's suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina HiSeq 2500,,SRP455680,,,Fish_embryo_mRNA_128cell_PAL_seq_v4_rep1_raw_read1.fastq.gz Fish_embryo_mRNA_128cell_PAL_seq_v4_rep1_raw_read2.fastq.gz,fastq fastq,2104868443.0,6856249.0,GSM7716867 r1,0:52 1:255,A:539122676;C:505234642;G:558793048;T:495876292;N:5841785,52,255,,,539122676,505234642,558793048,495876292,5841785,SRX21396038,SRS18636196,SRA1694849,Whitehead Institute,Whitehead Institute,2,0.00035,0.29379,7e-05,0.01129,0.99949,0.99971,0.55882,0.79591,52,255,T,B,mate1 technical by mapping diff,illumina,hiseq_era,unknown,poly_a,unknown,bulk,bulk,bulk,,United States,2023-08-17,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
25191,SRR25670738,SRX21396038,SRS18636196,SRP455680,PRJNA1006406,Control of polyA tail length and translation in vertebrate oocytes and early embryos,GSE241107,Other,During oocyte maturation and early embryonic development  polyA tail lengths strongly influence mRNA translation. However  how tail lengths are controlled at different developmental stages has been unclear. Here  we performed tail length and translational profiling of mRNA reporter libraries each with > 10 million three prime UTR sequence variants in frog oocytes and embryos  and fish embryos. These analyses revealed that the UUUUA motif specifies cytoplasmic polyadenylation and identified diverse context features that modulate the activity of this 5 mer. Additional sequence motifs drive stage specific deadenylation in embryos  and UUUUA and C rich motifs drive tail length independent translational repression in oocytes. A neural network model accurately predicts tail length change during oocyte maturation in frogs  mice  and humans. Analyses of human sequence variants showed that those predicted to disrupt tail length control have been under negative selection  implying that our insights into control of polyA tail length and translation have implications for human health and fertility. Overall design: Sythetic mRNA reporter libraries with random three prime UTR sequences under four different sequence contexts were injected into frog oocytes and embryos and fish embryos. PolyA tail lengths were measured to at different developmental stages to examine sequence motifs that caused tail length changes. At the same time  polyA tail lengths of endogenous mRNAs from frog oocytes and embryos  fish embryos  and mouse oocytes were measured to investiage how their tail lengths were controlled. Please note that sample titles have been updated on Jan 6  2024.,,,,Fish embryo mRNA 128cell PAL seq v4,GSM7716867,,source name:embryo|tissue:embryo|treatment:N1|geo loc name:missing|collection date:missing,Fish embryo mRNA 128cell PAL seq v4,For gene specific tail seq of mRNA reporters  data were processed with a custom script available at https://github.com/coffeebond/MPRA tail seq. For PAL seq v3 or v4  reads were trimmed with cutadapt v3.7 with the parameters “ m 15   quality base=64  q 20 20   match read wildcards  e 0.05  a NNNNATCTCGTATGCCGTCTTCTGCTTG  O 7”. The trimmed reads were mapped using STAR v2.7.1a to the reference database containing the genomic sequences of the organism from which the mRNAs were obtained  the genomic sequences of humans or fish  depending on which spike in RNAs were used  and the sequences of the polyA standards generated previously  with the parameters “  runThreadN 16   runMode alignReads   outFilterMultimapNmax 1   outReadsUnmapped Fastx   outFilterType BySJout   outSAMattributes All   outSAMtype BAM Unsorted SortedByCoordinate”. Uniquely mapped read were furhter processed with a custom script available at https://github.com/coffeebond/PAL seq. Assembly: Homo sapiens: GRCh38.p7 primary assembly. Mus muculus: GRCh38.p4  primary assembly. Xenopus laevis: v10.1 assembly. Danio rerio: GRCz11 assembly. Supplementary files format and content: For gene specific tail seq of mRNA reporters  tab delimited text files with columns indicating 1 sequence of the variable region; 2 median tail length; 3 number of reads; 4–7 tail lengths at quantile 10  25  75  and 90. Supplementary files format and content: For PAL seq v3 or v4  tab delimited files with columns indicating 1 gene ID or polyA site ID; 2 read cluster ID; 3 tail length Library strategy: PAL seq v4,embryo,,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer’s suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,tissue:embryo|treatment:N1,GSM7716867,GSM7716867: Fish embryo mRNA 128cell PAL seq v4; Danio rerio; OTHER,GSM7716867 r1,GSM7716867,1,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer's suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina HiSeq 2500,,SRP455680,,,Fish_embryo_mRNA_128cell_PAL_seq_v4_rep2_raw_read1.fastq.gz Fish_embryo_mRNA_128cell_PAL_seq_v4_rep2_raw_read2.fastq.gz,fastq fastq,3135806371.0,10214353.0,GSM7716867 r2,0:52 1:255,A:776612486;C:774126305;G:853951502;T:724788612;N:6327466,52,255,,,776612486,774126305,853951502,724788612,6327466,SRX21396038,SRS18636196,SRA1694849,Whitehead Institute,Whitehead Institute,2,0.00066,0.0,0.00011,0.0,0.99939,1.0,0.55737,,52,255,T,T,mates < 9% mapping rate,illumina,hiseq_era,unknown,poly_a,unknown,bulk,bulk,bulk,,United States,2023-08-17,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
25192,SRR25670739,SRX21396037,SRS18636195,SRP455680,PRJNA1006406,Control of polyA tail length and translation in vertebrate oocytes and early embryos,GSE241107,Other,During oocyte maturation and early embryonic development  polyA tail lengths strongly influence mRNA translation. However  how tail lengths are controlled at different developmental stages has been unclear. Here  we performed tail length and translational profiling of mRNA reporter libraries each with > 10 million three prime UTR sequence variants in frog oocytes and embryos  and fish embryos. These analyses revealed that the UUUUA motif specifies cytoplasmic polyadenylation and identified diverse context features that modulate the activity of this 5 mer. Additional sequence motifs drive stage specific deadenylation in embryos  and UUUUA and C rich motifs drive tail length independent translational repression in oocytes. A neural network model accurately predicts tail length change during oocyte maturation in frogs  mice  and humans. Analyses of human sequence variants showed that those predicted to disrupt tail length control have been under negative selection  implying that our insights into control of polyA tail length and translation have implications for human health and fertility. Overall design: Sythetic mRNA reporter libraries with random three prime UTR sequences under four different sequence contexts were injected into frog oocytes and embryos and fish embryos. PolyA tail lengths were measured to at different developmental stages to examine sequence motifs that caused tail length changes. At the same time  polyA tail lengths of endogenous mRNAs from frog oocytes and embryos  fish embryos  and mouse oocytes were measured to investiage how their tail lengths were controlled. Please note that sample titles have been updated on Jan 6  2024.,,,,Fish embryo mRNA 8cell PAL seq v4,GSM7716866,,source name:embryo|tissue:embryo|treatment:N1|geo loc name:missing|collection date:missing,Fish embryo mRNA 8cell PAL seq v4,For gene specific tail seq of mRNA reporters  data were processed with a custom script available at https://github.com/coffeebond/MPRA tail seq. For PAL seq v3 or v4  reads were trimmed with cutadapt v3.7 with the parameters “ m 15   quality base=64  q 20 20   match read wildcards  e 0.05  a NNNNATCTCGTATGCCGTCTTCTGCTTG  O 7”. The trimmed reads were mapped using STAR v2.7.1a to the reference database containing the genomic sequences of the organism from which the mRNAs were obtained  the genomic sequences of humans or fish  depending on which spike in RNAs were used  and the sequences of the polyA standards generated previously  with the parameters “  runThreadN 16   runMode alignReads   outFilterMultimapNmax 1   outReadsUnmapped Fastx   outFilterType BySJout   outSAMattributes All   outSAMtype BAM Unsorted SortedByCoordinate”. Uniquely mapped read were furhter processed with a custom script available at https://github.com/coffeebond/PAL seq. Assembly: Homo sapiens: GRCh38.p7 primary assembly. Mus muculus: GRCh38.p4  primary assembly. Xenopus laevis: v10.1 assembly. Danio rerio: GRCz11 assembly. Supplementary files format and content: For gene specific tail seq of mRNA reporters  tab delimited text files with columns indicating 1 sequence of the variable region; 2 median tail length; 3 number of reads; 4–7 tail lengths at quantile 10  25  75  and 90. Supplementary files format and content: For PAL seq v3 or v4  tab delimited files with columns indicating 1 gene ID or polyA site ID; 2 read cluster ID; 3 tail length Library strategy: PAL seq v4,embryo,,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer’s suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,tissue:embryo|treatment:N1,GSM7716866,GSM7716866: Fish embryo mRNA 8cell PAL seq v4; Danio rerio; OTHER,GSM7716866 r1,GSM7716866,1,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer's suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina HiSeq 2500,,SRP455680,,,Fish_embryo_mRNA_8cell_PAL_seq_v4_rep1_raw_read1.fastq.gz Fish_embryo_mRNA_8cell_PAL_seq_v4_rep1_raw_read2.fastq.gz,fastq fastq,2618998887.0,8530941.0,GSM7716866 r1,0:52 1:255,A:673066508;C:641446717;G:706546263;T:590452494;N:7486905,52,255,,,673066508,641446717,706546263,590452494,7486905,SRX21396037,SRS18636195,SRA1694849,Whitehead Institute,Whitehead Institute,2,0.0009,0.43244,0.00014,0.0054,0.99902,0.99967,0.54901,1.0,52,255,T,B,mate1 technical by mapping diff,illumina,hiseq_era,unknown,poly_a,unknown,bulk,bulk,bulk,,United States,2023-08-17,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
25193,SRR25670740,SRX21396037,SRS18636195,SRP455680,PRJNA1006406,Control of polyA tail length and translation in vertebrate oocytes and early embryos,GSE241107,Other,During oocyte maturation and early embryonic development  polyA tail lengths strongly influence mRNA translation. However  how tail lengths are controlled at different developmental stages has been unclear. Here  we performed tail length and translational profiling of mRNA reporter libraries each with > 10 million three prime UTR sequence variants in frog oocytes and embryos  and fish embryos. These analyses revealed that the UUUUA motif specifies cytoplasmic polyadenylation and identified diverse context features that modulate the activity of this 5 mer. Additional sequence motifs drive stage specific deadenylation in embryos  and UUUUA and C rich motifs drive tail length independent translational repression in oocytes. A neural network model accurately predicts tail length change during oocyte maturation in frogs  mice  and humans. Analyses of human sequence variants showed that those predicted to disrupt tail length control have been under negative selection  implying that our insights into control of polyA tail length and translation have implications for human health and fertility. Overall design: Sythetic mRNA reporter libraries with random three prime UTR sequences under four different sequence contexts were injected into frog oocytes and embryos and fish embryos. PolyA tail lengths were measured to at different developmental stages to examine sequence motifs that caused tail length changes. At the same time  polyA tail lengths of endogenous mRNAs from frog oocytes and embryos  fish embryos  and mouse oocytes were measured to investiage how their tail lengths were controlled. Please note that sample titles have been updated on Jan 6  2024.,,,,Fish embryo mRNA 8cell PAL seq v4,GSM7716866,,source name:embryo|tissue:embryo|treatment:N1|geo loc name:missing|collection date:missing,Fish embryo mRNA 8cell PAL seq v4,For gene specific tail seq of mRNA reporters  data were processed with a custom script available at https://github.com/coffeebond/MPRA tail seq. For PAL seq v3 or v4  reads were trimmed with cutadapt v3.7 with the parameters “ m 15   quality base=64  q 20 20   match read wildcards  e 0.05  a NNNNATCTCGTATGCCGTCTTCTGCTTG  O 7”. The trimmed reads were mapped using STAR v2.7.1a to the reference database containing the genomic sequences of the organism from which the mRNAs were obtained  the genomic sequences of humans or fish  depending on which spike in RNAs were used  and the sequences of the polyA standards generated previously  with the parameters “  runThreadN 16   runMode alignReads   outFilterMultimapNmax 1   outReadsUnmapped Fastx   outFilterType BySJout   outSAMattributes All   outSAMtype BAM Unsorted SortedByCoordinate”. Uniquely mapped read were furhter processed with a custom script available at https://github.com/coffeebond/PAL seq. Assembly: Homo sapiens: GRCh38.p7 primary assembly. Mus muculus: GRCh38.p4  primary assembly. Xenopus laevis: v10.1 assembly. Danio rerio: GRCz11 assembly. Supplementary files format and content: For gene specific tail seq of mRNA reporters  tab delimited text files with columns indicating 1 sequence of the variable region; 2 median tail length; 3 number of reads; 4–7 tail lengths at quantile 10  25  75  and 90. Supplementary files format and content: For PAL seq v3 or v4  tab delimited files with columns indicating 1 gene ID or polyA site ID; 2 read cluster ID; 3 tail length Library strategy: PAL seq v4,embryo,,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer’s suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,tissue:embryo|treatment:N1,GSM7716866,GSM7716866: Fish embryo mRNA 8cell PAL seq v4; Danio rerio; OTHER,GSM7716866 r1,GSM7716866,1,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer's suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina HiSeq 2500,,SRP455680,,,Fish_embryo_mRNA_8cell_PAL_seq_v4_rep2_raw_read1.fastq.gz Fish_embryo_mRNA_8cell_PAL_seq_v4_rep2_raw_read2.fastq.gz,fastq fastq,3028839589.0,9865927.0,GSM7716866 r2,0:52 1:255,A:756368817;C:758868409;G:836949742;T:670545278;N:6107343,52,255,,,756368817,758868409,836949742,670545278,6107343,SRX21396037,SRS18636195,SRA1694849,Whitehead Institute,Whitehead Institute,2,0.00156,0.0,0.00026,0.0,0.99835,1.0,0.44791,,52,255,T,T,mates < 9% mapping rate,illumina,hiseq_era,unknown,poly_a,unknown,bulk,bulk,bulk,,United States,2023-08-17,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
25194,SRR25670741,SRX21396036,SRS18636194,SRP455680,PRJNA1006406,Control of polyA tail length and translation in vertebrate oocytes and early embryos,GSE241107,Other,During oocyte maturation and early embryonic development  polyA tail lengths strongly influence mRNA translation. However  how tail lengths are controlled at different developmental stages has been unclear. Here  we performed tail length and translational profiling of mRNA reporter libraries each with > 10 million three prime UTR sequence variants in frog oocytes and embryos  and fish embryos. These analyses revealed that the UUUUA motif specifies cytoplasmic polyadenylation and identified diverse context features that modulate the activity of this 5 mer. Additional sequence motifs drive stage specific deadenylation in embryos  and UUUUA and C rich motifs drive tail length independent translational repression in oocytes. A neural network model accurately predicts tail length change during oocyte maturation in frogs  mice  and humans. Analyses of human sequence variants showed that those predicted to disrupt tail length control have been under negative selection  implying that our insights into control of polyA tail length and translation have implications for human health and fertility. Overall design: Sythetic mRNA reporter libraries with random three prime UTR sequences under four different sequence contexts were injected into frog oocytes and embryos and fish embryos. PolyA tail lengths were measured to at different developmental stages to examine sequence motifs that caused tail length changes. At the same time  polyA tail lengths of endogenous mRNAs from frog oocytes and embryos  fish embryos  and mouse oocytes were measured to investiage how their tail lengths were controlled. Please note that sample titles have been updated on Jan 6  2024.,,,,Fish embryo mRNA 1cell PAL seq v4,GSM7716865,,source name:embryo|tissue:embryo|treatment:N1|geo loc name:missing|collection date:missing,Fish embryo mRNA 1cell PAL seq v4,For gene specific tail seq of mRNA reporters  data were processed with a custom script available at https://github.com/coffeebond/MPRA tail seq. For PAL seq v3 or v4  reads were trimmed with cutadapt v3.7 with the parameters “ m 15   quality base=64  q 20 20   match read wildcards  e 0.05  a NNNNATCTCGTATGCCGTCTTCTGCTTG  O 7”. The trimmed reads were mapped using STAR v2.7.1a to the reference database containing the genomic sequences of the organism from which the mRNAs were obtained  the genomic sequences of humans or fish  depending on which spike in RNAs were used  and the sequences of the polyA standards generated previously  with the parameters “  runThreadN 16   runMode alignReads   outFilterMultimapNmax 1   outReadsUnmapped Fastx   outFilterType BySJout   outSAMattributes All   outSAMtype BAM Unsorted SortedByCoordinate”. Uniquely mapped read were furhter processed with a custom script available at https://github.com/coffeebond/PAL seq. Assembly: Homo sapiens: GRCh38.p7 primary assembly. Mus muculus: GRCh38.p4  primary assembly. Xenopus laevis: v10.1 assembly. Danio rerio: GRCz11 assembly. Supplementary files format and content: For gene specific tail seq of mRNA reporters  tab delimited text files with columns indicating 1 sequence of the variable region; 2 median tail length; 3 number of reads; 4–7 tail lengths at quantile 10  25  75  and 90. Supplementary files format and content: For PAL seq v3 or v4  tab delimited files with columns indicating 1 gene ID or polyA site ID; 2 read cluster ID; 3 tail length Library strategy: PAL seq v4,embryo,,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer’s suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,tissue:embryo|treatment:N1,GSM7716865,GSM7716865: Fish embryo mRNA 1cell PAL seq v4; Danio rerio; OTHER,GSM7716865 r1,GSM7716865,1,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer's suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina HiSeq 2500,,SRP455680,,,Fish_embryo_mRNA_1cell_PAL_seq_v4_rep1_raw_read1.fastq.gz Fish_embryo_mRNA_1cell_PAL_seq_v4_rep1_raw_read2.fastq.gz,fastq fastq,2050143851.0,6677993.0,GSM7716865 r1,0:52 1:255,A:536391564;C:527126186;G:557802929;T:422990166;N:5833006,52,255,,,536391564,527126186,557802929,422990166,5833006,SRX21396036,SRS18636194,SRA1694849,Whitehead Institute,Whitehead Institute,2,0.00016,0.46479,4e-05,0.01408,0.99979,0.99969,0.54545,1.0,52,255,T,B,mate1 technical by mapping diff,illumina,hiseq_era,unknown,poly_a,unknown,bulk,bulk,bulk,,United States,2023-08-17,Zygote,Embryo,Embryo Imprecise,All anatomical structures
25195,SRR25670742,SRX21396036,SRS18636194,SRP455680,PRJNA1006406,Control of polyA tail length and translation in vertebrate oocytes and early embryos,GSE241107,Other,During oocyte maturation and early embryonic development  polyA tail lengths strongly influence mRNA translation. However  how tail lengths are controlled at different developmental stages has been unclear. Here  we performed tail length and translational profiling of mRNA reporter libraries each with > 10 million three prime UTR sequence variants in frog oocytes and embryos  and fish embryos. These analyses revealed that the UUUUA motif specifies cytoplasmic polyadenylation and identified diverse context features that modulate the activity of this 5 mer. Additional sequence motifs drive stage specific deadenylation in embryos  and UUUUA and C rich motifs drive tail length independent translational repression in oocytes. A neural network model accurately predicts tail length change during oocyte maturation in frogs  mice  and humans. Analyses of human sequence variants showed that those predicted to disrupt tail length control have been under negative selection  implying that our insights into control of polyA tail length and translation have implications for human health and fertility. Overall design: Sythetic mRNA reporter libraries with random three prime UTR sequences under four different sequence contexts were injected into frog oocytes and embryos and fish embryos. PolyA tail lengths were measured to at different developmental stages to examine sequence motifs that caused tail length changes. At the same time  polyA tail lengths of endogenous mRNAs from frog oocytes and embryos  fish embryos  and mouse oocytes were measured to investiage how their tail lengths were controlled. Please note that sample titles have been updated on Jan 6  2024.,,,,Fish embryo mRNA 1cell PAL seq v4,GSM7716865,,source name:embryo|tissue:embryo|treatment:N1|geo loc name:missing|collection date:missing,Fish embryo mRNA 1cell PAL seq v4,For gene specific tail seq of mRNA reporters  data were processed with a custom script available at https://github.com/coffeebond/MPRA tail seq. For PAL seq v3 or v4  reads were trimmed with cutadapt v3.7 with the parameters “ m 15   quality base=64  q 20 20   match read wildcards  e 0.05  a NNNNATCTCGTATGCCGTCTTCTGCTTG  O 7”. The trimmed reads were mapped using STAR v2.7.1a to the reference database containing the genomic sequences of the organism from which the mRNAs were obtained  the genomic sequences of humans or fish  depending on which spike in RNAs were used  and the sequences of the polyA standards generated previously  with the parameters “  runThreadN 16   runMode alignReads   outFilterMultimapNmax 1   outReadsUnmapped Fastx   outFilterType BySJout   outSAMattributes All   outSAMtype BAM Unsorted SortedByCoordinate”. Uniquely mapped read were furhter processed with a custom script available at https://github.com/coffeebond/PAL seq. Assembly: Homo sapiens: GRCh38.p7 primary assembly. Mus muculus: GRCh38.p4  primary assembly. Xenopus laevis: v10.1 assembly. Danio rerio: GRCz11 assembly. Supplementary files format and content: For gene specific tail seq of mRNA reporters  tab delimited text files with columns indicating 1 sequence of the variable region; 2 median tail length; 3 number of reads; 4–7 tail lengths at quantile 10  25  75  and 90. Supplementary files format and content: For PAL seq v3 or v4  tab delimited files with columns indicating 1 gene ID or polyA site ID; 2 read cluster ID; 3 tail length Library strategy: PAL seq v4,embryo,,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer’s suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,tissue:embryo|treatment:N1,GSM7716865,GSM7716865: Fish embryo mRNA 1cell PAL seq v4; Danio rerio; OTHER,GSM7716865 r1,GSM7716865,1,Frog oocytes and embryos were lysed in ice cold buffer RL 20 mM HEPES pH 7.5  100 mM KCl  5 mM MgCl2  1% [v/v] Triton X 100  100 µg/ml cycloheximide  cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer]  and 200 units/ml SUPERase•In in a volume of 10 µl per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4°C for 10 min. The supernatant was transfered to a new tube and mixed with the Tri reagent for RNA isolation. Fish embryos were de chorionated by incubation with 2 mg/ml pronase in E3 medium for 4 min. post removing all E3 medium  Tri Reagent was added to the embryos for RNA isolation. Mouse GV oocytes were collected in 37°C MEM in the presence of milrinone to prevent maturation. Cumulus cells were removed from cumulus oocyte complexes by repeated aspiration through a glass pipette. GV oocytes were then collected in TRI reagent for RNA isolation. Mouse MII oocytes were harvested from the oviducts of hCG induced mice 16 hr  denuded with 3 mg/ml hyaluronidase for 2 min  washed  and then collected in Tri reagent for RNA isolation. For gene specific tail seq of reporter libraries  total RNA with reporter mRNA libraries was ligated to a pre adenylated 3ʹ adapter directly in most cases  but for the N60 library injected into fish embryos and frog oocytes  the N37 PAS N17 library injected into fish embryos and frog oocytes  and the CPEmos N60 and N60LC PASmos libraries injected into frog oocytes  reporter library mRNAs were enriched by anti sense oligo capture with biotinylated oligos. RNA isolated from the oocyte or embryo lysate was mixed with 8 pmol KXSH009  8 pmol KXSH010  and 2x SSC 0.3 M NaCl  30 mM sodium citrate pH 7.0 in a total of 50 µl. The RNA and oligos were annealed by incubation at 70°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 40 µl MyOne Streptavidin C1 beads Thermo Fisher  65002 and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack and removed. The beads were washed twice with 300 µl 1xB&W buffer 5 mM Tris HCl pH 7.5  0.5 mM EDTA  1 M NaCl and once with 300 µl 2x SSC. The RNA was eluted from the beads first with 100 µl 10 mM HEPES pH 7.5 at 65°C for 3 min and then second with 100 µl water at 65°C for 3 min. The eluates were combined  precipitated with ethanol  and resuspended in 6.5 µl water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3ʹ adapter in a 10 µl reaction containing 5 µM 3ʹ adapter KXS330  50 mM HEPES pH 7.5  10 mM MgCl2  10 mM dithiothreitol  1 unit/µl T4 RNA ligase 1 New England Biolabs  M0204S. The ligation reaction was incubated at 23°C for 150 min. post ligation  RNA was extracted with phenol/chloroform  precipitated with ethanol  and resuspended in 11.4 µl water. The ligated RNA was mixed with 0.6 µl 100 µM reverse transcription primer KXS037 in a total volume of 12 µl  incubated at 65°C for 5 min  and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 µl reaction containing 1x First Strand Buffer  500 µM dNTPs  5 mM dithiothreitol  1 unit/µl SUPERase•In  and 200 units SuperScript III Thermo Fisher  18080044 at 50°C for 1 hr. post reverse transcription  RNA was hydrolyzed with 3.3 µl 1 M NaOH at 90°C for 10 min  followed by neutralization with 36.7 µl 1 M HEPES pH 7.5 and the cDNA was collected by desalting with a Micro Bio Spin P 30 column. The cDNA library was amplified in a 50 µl PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer's suggested protocol for 10–15 cycles. The PCR amplified library was cleaned up twice with AMPure XP beads Beckman Coulter  A63881 with a beads to sample ratio of 1.2. For sequencing of endogenous mRNA polyA tail length  libraries were prepared with PAL seq v3 for frog oocytes or PAL seq v4 for frog embryos  fish embryos  and mouse oocytes as described previously PMID: 34213414. When preparing the sequencing libraries of mRNAs from fish embryos  a different 3ʹ adapter KXS013 was used for 3ʹ end ligation  and polyA selected mRNA from HeLa cells was used as spike in  replacing polyA selected mRNA from zebrafish ZF4 cell line. The first round of sequencing results suggested that a large fraction of the fish mRNA libraries was 5.8S rRNA. The cDNA of 5.8S rRNA was depleted from the cDNA libraries with an antisense oligo. The seven cDNA libraries made from mRNAs of zebrafish embryos at different stages were mixed at roughly equal molar ratios in a total of 5 fmol. Fifty pmol KXSH015 and 2x SSC were added in a total of 100 µl. The cDNAs and the oligo were annealed by incubation at 65°C for 5 min and then slowly cooling to 23°C at 0.1°C/sec. The annealed mixture was combined with 100 µl MyOne Streptavidin C1 beads and incubated for 20 min at 23°C on a thermal mixer  shaking with 15 sec on and 1 min 45 sec off. The supernatant was separated from the beads with a magnetic rack. The beads were washed once with 200 µl 1xB&W buffer. The supernatant and the wash were combined  precipitated with ethanol  and resuspended in 30 µl water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina HiSeq 2500,,SRP455680,,,Fish_embryo_mRNA_1cell_PAL_seq_v4_rep2_raw_read1.fastq.gz Fish_embryo_mRNA_1cell_PAL_seq_v4_rep2_raw_read2.fastq.gz,fastq fastq,3406093776.0,11094768.0,GSM7716865 r2,0:52 1:255,A:868054279;C:890446070;G:945289336;T:695324057;N:6980034,52,255,,,868054279,890446070,945289336,695324057,6980034,SRX21396036,SRS18636194,SRA1694849,Whitehead Institute,Whitehead Institute,2,0.00049,0.0,0.00014,0.0,0.99939,1.0,0.58974,,52,255,T,T,mates < 9% mapping rate,illumina,hiseq_era,unknown,poly_a,unknown,bulk,bulk,bulk,,United States,2023-08-17,Zygote,Embryo,Embryo Imprecise,All anatomical structures
33990,SRR31030860,SRX26416598,SRS22936450,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF NES GAPDH Rep4 minus,GSM8578751,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF|geo loc name:missing|collection date:missing,ZF NES GAPDH Rep4 minus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF,GSM8578751,GSM8578751: ZF NES GAPDH Rep4 minus; Danio rerio; OTHER,GSM8578751 r1,GSM8578751,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_NES_GAPDH_Rep4_minus.R1.fq.gz ZF_NES_GAPDH_Rep4_minus.R2.fq.gz,fastq fastq,1100578200.0,3668594.0,GSM8578751 r1,0:150 1:150,A:287878221;C:249390946;G:265835736;T:297454600;N:18697,150,150,,,287878221,249390946,265835736,297454600,18697,SRX26416598,SRS22936450,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
33991,SRR31030861,SRX26416597,SRS22936451,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF NES GAPDH Rep3 plus,GSM8578750,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF|geo loc name:missing|collection date:missing,ZF NES GAPDH Rep3 plus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF,GSM8578750,GSM8578750: ZF NES GAPDH Rep3 plus; Danio rerio; OTHER,GSM8578750 r1,GSM8578750,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_NES_GAPDH_Rep3_plus.R1.fq.gz ZF_NES_GAPDH_Rep3_plus.R2.fq.gz,fastq fastq,1231779300.0,4105931.0,GSM8578750 r1,0:150 1:150,A:322256597;C:278991471;G:297394980;T:333116282;N:19970,150,150,,,322256597,278991471,297394980,333116282,19970,SRX26416597,SRS22936451,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
33992,SRR31030862,SRX26416596,SRS22936449,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF NES GAPDH Rep3 minus,GSM8578749,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF|geo loc name:missing|collection date:missing,ZF NES GAPDH Rep3 minus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF,GSM8578749,GSM8578749: ZF NES GAPDH Rep3 minus; Danio rerio; OTHER,GSM8578749 r1,GSM8578749,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_NES_GAPDH_Rep3_minus.R1.fq.gz ZF_NES_GAPDH_Rep3_minus.R2.fq.gz,fastq fastq,1002124500.0,3340415.0,GSM8578749 r1,0:150 1:150,A:262024969;C:227163761;G:242157218;T:270761424;N:17128,150,150,,,262024969,227163761,242157218,270761424,17128,SRX26416596,SRS22936449,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
33993,SRR31030863,SRX26416595,SRS22936448,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF NES GAPDH Rep2 plus,GSM8578748,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF|geo loc name:missing|collection date:missing,ZF NES GAPDH Rep2 plus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF,GSM8578748,GSM8578748: ZF NES GAPDH Rep2 plus; Danio rerio; OTHER,GSM8578748 r1,GSM8578748,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_NES_GAPDH_Rep2_plus.R1.fq.gz ZF_NES_GAPDH_Rep2_plus.R2.fq.gz,fastq fastq,1051565100.0,3505217.0,GSM8578748 r1,0:150 1:150,A:275784945;C:237523425;G:253300749;T:284937948;N:18033,150,150,,,275784945,237523425,253300749,284937948,18033,SRX26416595,SRS22936448,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
33994,SRR31030864,SRX26416594,SRS22936447,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF NES GAPDH Rep2 minus,GSM8578747,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF|geo loc name:missing|collection date:missing,ZF NES GAPDH Rep2 minus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF,GSM8578747,GSM8578747: ZF NES GAPDH Rep2 minus; Danio rerio; OTHER,GSM8578747 r1,GSM8578747,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_NES_GAPDH_Rep2_minus.R1.fq.gz ZF_NES_GAPDH_Rep2_minus.R2.fq.gz,fastq fastq,1021837200.0,3406124.0,GSM8578747 r1,0:150 1:150,A:268290250;C:230525208;G:245949056;T:277056045;N:16641,150,150,,,268290250,230525208,245949056,277056045,16641,SRX26416594,SRS22936447,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
33995,SRR31030865,SRX26416593,SRS22936446,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF NES GAPDH Rep1 plus,GSM8578746,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF|geo loc name:missing|collection date:missing,ZF NES GAPDH Rep1 plus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF,GSM8578746,GSM8578746: ZF NES GAPDH Rep1 plus; Danio rerio; OTHER,GSM8578746 r1,GSM8578746,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_NES_GAPDH_Rep1_plus.R1.fq.gz ZF_NES_GAPDH_Rep1_plus.R2.fq.gz,fastq fastq,1093620300.0,3645401.0,GSM8578746 r1,0:150 1:150,A:285989719;C:247805250;G:264181043;T:295625946;N:18342,150,150,,,285989719,247805250,264181043,295625946,18342,SRX26416593,SRS22936446,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
33996,SRR31030866,SRX26416592,SRS22936445,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF NES GAPDH Rep1 minus,GSM8578745,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF|geo loc name:missing|collection date:missing,ZF NES GAPDH Rep1 minus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF,GSM8578745,GSM8578745: ZF NES GAPDH Rep1 minus; Danio rerio; OTHER,GSM8578745 r1,GSM8578745,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_NES_GAPDH_Rep1_minus.R1.fq.gz ZF_NES_GAPDH_Rep1_minus.R2.fq.gz,fastq fastq,911747100.0,3039157.0,GSM8578745 r1,0:150 1:150,A:239833349;C:205185786;G:218897710;T:247814511;N:15744,150,150,,,239833349,205185786,218897710,247814511,15744,SRX26416592,SRS22936445,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
33997,SRR31030867,SRX26416591,SRS22936444,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF H2B MALAT1 Rep4 plus,GSM8578744,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo H2B|treatment:plus DBF|geo loc name:missing|collection date:missing,ZF H2B MALAT1 Rep4 plus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo H2B|treatment:plus DBF,GSM8578744,GSM8578744: ZF H2B MALAT1 Rep4 plus; Danio rerio; OTHER,GSM8578744 r1,GSM8578744,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_H2B_MALAT1_Rep4_plus.R1.fq.gz ZF_H2B_MALAT1_Rep4_plus.R2.fq.gz,fastq fastq,842103300.0,2807011.0,GSM8578744 r1,0:150 1:150,A:204870477;C:222752099;G:215404135;T:199062275;N:14314,150,150,,,204870477,222752099,215404135,199062275,14314,SRX26416591,SRS22936444,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
33998,SRR31030868,SRX26416590,SRS22936442,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF H2B MALAT1 Rep4 minus,GSM8578743,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo H2B|treatment:minus DBF|geo loc name:missing|collection date:missing,ZF H2B MALAT1 Rep4 minus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo H2B|treatment:minus DBF,GSM8578743,GSM8578743: ZF H2B MALAT1 Rep4 minus; Danio rerio; OTHER,GSM8578743 r1,GSM8578743,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_H2B_MALAT1_Rep4_minus.R1.fq.gz ZF_H2B_MALAT1_Rep4_minus.R2.fq.gz,fastq fastq,1166375100.0,3887917.0,GSM8578743 r1,0:150 1:150,A:283781955;C:308660466;G:298298204;T:275614377;N:20098,150,150,,,283781955,308660466,298298204,275614377,20098,SRX26416590,SRS22936442,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
33999,SRR31030869,SRX26416589,SRS22936443,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF H2B MALAT1 Rep3 plus,GSM8578742,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo H2B|treatment:plus DBF|geo loc name:missing|collection date:missing,ZF H2B MALAT1 Rep3 plus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo H2B|treatment:plus DBF,GSM8578742,GSM8578742: ZF H2B MALAT1 Rep3 plus; Danio rerio; OTHER,GSM8578742 r1,GSM8578742,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_H2B_MALAT1_Rep3_plus.R1.fq.gz ZF_H2B_MALAT1_Rep3_plus.R2.fq.gz,fastq fastq,1159439400.0,3864798.0,GSM8578742 r1,0:150 1:150,A:282055391;C:306842216;G:296569227;T:273952570;N:19996,150,150,,,282055391,306842216,296569227,273952570,19996,SRX26416589,SRS22936443,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34000,SRR31030870,SRX26416588,SRS22936440,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF H2B MALAT1 Rep3 minus,GSM8578741,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo H2B|treatment:minus DBF|geo loc name:missing|collection date:missing,ZF H2B MALAT1 Rep3 minus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo H2B|treatment:minus DBF,GSM8578741,GSM8578741: ZF H2B MALAT1 Rep3 minus; Danio rerio; OTHER,GSM8578741 r1,GSM8578741,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_H2B_MALAT1_Rep3_minus.R1.fq.gz ZF_H2B_MALAT1_Rep3_minus.R2.fq.gz,fastq fastq,1008297600.0,3360992.0,GSM8578741 r1,0:150 1:150,A:245277585;C:266840408;G:257911846;T:238251305;N:16456,150,150,,,245277585,266840408,257911846,238251305,16456,SRX26416588,SRS22936440,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34001,SRR31030871,SRX26416587,SRS22936441,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF H2B MALAT1 Rep2 plus,GSM8578740,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo H2B|treatment:plus DBF|geo loc name:missing|collection date:missing,ZF H2B MALAT1 Rep2 plus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo H2B|treatment:plus DBF,GSM8578740,GSM8578740: ZF H2B MALAT1 Rep2 plus; Danio rerio; OTHER,GSM8578740 r1,GSM8578740,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_H2B_MALAT1_Rep2_plus.R1.fq.gz ZF_H2B_MALAT1_Rep2_plus.R2.fq.gz,fastq fastq,1419263400.0,4730878.0,GSM8578740 r1,0:150 1:150,A:345302268;C:375591885;G:362956395;T:335387828;N:25024,150,150,,,345302268,375591885,362956395,335387828,25024,SRX26416587,SRS22936441,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34002,SRR31030872,SRX26416586,SRS22936439,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF H2B MALAT1 Rep2 minus,GSM8578739,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo H2B|treatment:minus DBF|geo loc name:missing|collection date:missing,ZF H2B MALAT1 Rep2 minus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo H2B|treatment:minus DBF,GSM8578739,GSM8578739: ZF H2B MALAT1 Rep2 minus; Danio rerio; OTHER,GSM8578739 r1,GSM8578739,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_H2B_MALAT1_Rep2_minus.R1.fq.gz ZF_H2B_MALAT1_Rep2_minus.R2.fq.gz,fastq fastq,1249910700.0,4166369.0,GSM8578739 r1,0:150 1:150,A:304074828;C:330741431;G:319677748;T:295395235;N:21458,150,150,,,304074828,330741431,319677748,295395235,21458,SRX26416586,SRS22936439,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34003,SRR31030873,SRX26416585,SRS22936438,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF H2B MALAT1 Rep1 plus,GSM8578738,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo H2B|treatment:plus DBF|geo loc name:missing|collection date:missing,ZF H2B MALAT1 Rep1 plus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo H2B|treatment:plus DBF,GSM8578738,GSM8578738: ZF H2B MALAT1 Rep1 plus; Danio rerio; OTHER,GSM8578738 r1,GSM8578738,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_H2B_MALAT1_Rep1_plus.R1.fq.gz ZF_H2B_MALAT1_Rep1_plus.R2.fq.gz,fastq fastq,1257872700.0,4192909.0,GSM8578738 r1,0:150 1:150,A:306022947;C:332851039;G:321699513;T:297277788;N:21413,150,150,,,306022947,332851039,321699513,297277788,21413,SRX26416585,SRS22936438,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34004,SRR31030874,SRX26416584,SRS22936437,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF H2B MALAT1 Rep1 minus,GSM8578737,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo H2B|treatment:minus DBF|geo loc name:missing|collection date:missing,ZF H2B MALAT1 Rep1 minus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo H2B|treatment:minus DBF,GSM8578737,GSM8578737: ZF H2B MALAT1 Rep1 minus; Danio rerio; OTHER,GSM8578737 r1,GSM8578737,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_H2B_MALAT1_Rep1_minus.R1.fq.gz ZF_H2B_MALAT1_Rep1_minus.R2.fq.gz,fastq fastq,1377040200.0,4590134.0,GSM8578737 r1,0:150 1:150,A:335021330;C:364359707;G:352178356;T:325457746;N:23061,150,150,,,335021330,364359707,352178356,325457746,23061,SRX26416584,SRS22936437,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34005,SRR31030875,SRX26416583,SRS22936435,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF H2B GAPDH Rep4 plus,GSM8578736,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo H2B|treatment:plus DBF|geo loc name:missing|collection date:missing,ZF H2B GAPDH Rep4 plus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo H2B|treatment:plus DBF,GSM8578736,GSM8578736: ZF H2B GAPDH Rep4 plus; Danio rerio; OTHER,GSM8578736 r1,GSM8578736,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_H2B_GAPDH_Rep4_plus.R1.fq.gz ZF_H2B_GAPDH_Rep4_plus.R2.fq.gz,fastq fastq,1051103100.0,3503677.0,GSM8578736 r1,0:150 1:150,A:274220771;C:238733668;G:254670769;T:283460283;N:17609,150,150,,,274220771,238733668,254670769,283460283,17609,SRX26416583,SRS22936435,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,mate1-mate2 similar by mapping diff,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34006,SRR31030876,SRX26416582,SRS22936436,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF H2B GAPDH Rep4 minus,GSM8578735,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo H2B|treatment:minus DBF|geo loc name:missing|collection date:missing,ZF H2B GAPDH Rep4 minus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo H2B|treatment:minus DBF,GSM8578735,GSM8578735: ZF H2B GAPDH Rep4 minus; Danio rerio; OTHER,GSM8578735 r1,GSM8578735,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_H2B_GAPDH_Rep4_minus.R1.fq.gz ZF_H2B_GAPDH_Rep4_minus.R2.fq.gz,fastq fastq,1206485700.0,4021619.0,GSM8578735 r1,0:150 1:150,A:314740194;C:274101014;G:292357972;T:325267132;N:19388,150,150,,,314740194,274101014,292357972,325267132,19388,SRX26416582,SRS22936436,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,mate1-mate2 similar by mapping diff,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34007,SRR31030877,SRX26416581,SRS22936434,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF H2B GAPDH Rep3 plus,GSM8578734,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo H2B|treatment:plus DBF|geo loc name:missing|collection date:missing,ZF H2B GAPDH Rep3 plus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo H2B|treatment:plus DBF,GSM8578734,GSM8578734: ZF H2B GAPDH Rep3 plus; Danio rerio; OTHER,GSM8578734 r1,GSM8578734,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_H2B_GAPDH_Rep3_plus.R1.fq.gz ZF_H2B_GAPDH_Rep3_plus.R2.fq.gz,fastq fastq,1140550200.0,3801834.0,GSM8578734 r1,0:150 1:150,A:297636297;C:258828826;G:276373993;T:307692213;N:18871,150,150,,,297636297,258828826,276373993,307692213,18871,SRX26416581,SRS22936434,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,mate1-mate2 similar by mapping diff,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34008,SRR31030878,SRX26416580,SRS22936433,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF H2B GAPDH Rep3 minus,GSM8578733,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo H2B|treatment:minus DBF|geo loc name:missing|collection date:missing,ZF H2B GAPDH Rep3 minus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo H2B|treatment:minus DBF,GSM8578733,GSM8578733: ZF H2B GAPDH Rep3 minus; Danio rerio; OTHER,GSM8578733 r1,GSM8578733,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_H2B_GAPDH_Rep3_minus.R1.fq.gz ZF_H2B_GAPDH_Rep3_minus.R2.fq.gz,fastq fastq,782406600.0,2608022.0,GSM8578733 r1,0:150 1:150,A:204132316;C:177696721;G:189675433;T:210888939;N:13191,150,150,,,204132316,177696721,189675433,210888939,13191,SRX26416580,SRS22936433,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,mate1-mate2 similar by mapping diff,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34009,SRR31030879,SRX26416579,SRS22936431,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF H2B GAPDH Rep2 plus,GSM8578732,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo H2B|treatment:plus DBF|geo loc name:missing|collection date:missing,ZF H2B GAPDH Rep2 plus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo H2B|treatment:plus DBF,GSM8578732,GSM8578732: ZF H2B GAPDH Rep2 plus; Danio rerio; OTHER,GSM8578732 r1,GSM8578732,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_H2B_GAPDH_Rep2_plus.R1.fq.gz ZF_H2B_GAPDH_Rep2_plus.R2.fq.gz,fastq fastq,958297200.0,3194324.0,GSM8578732 r1,0:150 1:150,A:250011335;C:217600823;G:232206339;T:258462392;N:16311,150,150,,,250011335,217600823,232206339,258462392,16311,SRX26416579,SRS22936431,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,mate1-mate2 similar by mapping diff,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34010,SRR31030880,SRX26416578,SRS22936432,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF H2B GAPDH Rep2 minus,GSM8578731,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo H2B|treatment:minus DBF|geo loc name:missing|collection date:missing,ZF H2B GAPDH Rep2 minus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo H2B|treatment:minus DBF,GSM8578731,GSM8578731: ZF H2B GAPDH Rep2 minus; Danio rerio; OTHER,GSM8578731 r1,GSM8578731,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_H2B_GAPDH_Rep2_minus.R1.fq.gz ZF_H2B_GAPDH_Rep2_minus.R2.fq.gz,fastq fastq,1020926700.0,3403089.0,GSM8578731 r1,0:150 1:150,A:266429488;C:231749424;G:247353147;T:275377720;N:16921,150,150,,,266429488,231749424,247353147,275377720,16921,SRX26416578,SRS22936432,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,mate1-mate2 similar by mapping diff,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34011,SRR31030881,SRX26416577,SRS22936430,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF H2B GAPDH Rep1 plus,GSM8578730,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo H2B|treatment:plus DBF|geo loc name:missing|collection date:missing,ZF H2B GAPDH Rep1 plus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo H2B|treatment:plus DBF,GSM8578730,GSM8578730: ZF H2B GAPDH Rep1 plus; Danio rerio; OTHER,GSM8578730 r1,GSM8578730,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_H2B_GAPDH_Rep1_plus.R1.fq.gz ZF_H2B_GAPDH_Rep1_plus.R2.fq.gz,fastq fastq,1134318600.0,3781062.0,GSM8578730 r1,0:150 1:150,A:295931335;C:257596481;G:274848790;T:305922827;N:19167,150,150,,,295931335,257596481,274848790,305922827,19167,SRX26416577,SRS22936430,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,mate1-mate2 similar by mapping diff,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34012,SRR31030882,SRX26416576,SRS22936429,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF H2B GAPDH Rep1 minus,GSM8578729,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo H2B|treatment:minus DBF|geo loc name:missing|collection date:missing,ZF H2B GAPDH Rep1 minus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo H2B|treatment:minus DBF,GSM8578729,GSM8578729: ZF H2B GAPDH Rep1 minus; Danio rerio; OTHER,GSM8578729 r1,GSM8578729,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_H2B_GAPDH_Rep1_minus.R1.fq.gz ZF_H2B_GAPDH_Rep1_minus.R2.fq.gz,fastq fastq,752638800.0,2508796.0,GSM8578729 r1,0:150 1:150,A:196331185;C:170899863;G:182460258;T:202935049;N:12445,150,150,,,196331185,170899863,182460258,202935049,12445,SRX26416576,SRS22936429,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,mate1-mate2 similar by mapping diff,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34013,SRR31031004,SRX26416454,SRS22936307,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,Zeb WT pDBF Rep3,GSM8578770,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:wildtype|treatment:plus DBF|geo loc name:missing|collection date:missing,Zeb WT pDBF Rep3,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:wildtype|treatment:plus DBF,GSM8578770,GSM8578770: Zeb WT pDBF Rep3; Danio rerio; OTHER,GSM8578770 r1,GSM8578770,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,Zeb_WT_pDBF_Rep3.R1.fq.gz Zeb_WT_pDBF_Rep3.R2.fq.gz,fastq fastq,23216831100.0,77389437.0,GSM8578770 r1,0:150 1:150,A:7577896837;C:3508911439;G:4823311862;T:7306393244;N:317718,150,150,,,7577896837,3508911439,4823311862,7306393244,317718,SRX26416454,SRS22936307,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,mate2-mate1 similar by mapping diff,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34014,SRR31031005,SRX26416453,SRS22936305,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,Zeb WT pDBF Rep2,GSM8578769,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:wildtype|treatment:plus DBF|geo loc name:missing|collection date:missing,Zeb WT pDBF Rep2,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:wildtype|treatment:plus DBF,GSM8578769,GSM8578769: Zeb WT pDBF Rep2; Danio rerio; OTHER,GSM8578769 r1,GSM8578769,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,Zeb_WT_pDBF_Rep2.R1.fq.gz Zeb_WT_pDBF_Rep2.R2.fq.gz,fastq fastq,27237049500.0,90790165.0,GSM8578769 r1,0:150 1:150,A:8800724632;C:4062692793;G:5651432074;T:8721823175;N:376826,150,150,,,8800724632,4062692793,5651432074,8721823175,376826,SRX26416453,SRS22936305,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,mate2-mate1 similar by mapping diff,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34015,SRR31031006,SRX26416452,SRS22936306,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,Zeb WT pDBF Rep1,GSM8578768,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:wildtype|treatment:plus DBF|geo loc name:missing|collection date:missing,Zeb WT pDBF Rep1,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:wildtype|treatment:plus DBF,GSM8578768,GSM8578768: Zeb WT pDBF Rep1; Danio rerio; OTHER,GSM8578768 r1,GSM8578768,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,Zeb_WT_pDBF_Rep1.R1.fq.gz Zeb_WT_pDBF_Rep1.R2.fq.gz,fastq fastq,22364115300.0,74547051.0,GSM8578768 r1,0:150 1:150,A:7336106577;C:3346831944;G:4451536371;T:7229335436;N:304972,150,150,,,7336106577,3346831944,4451536371,7229335436,304972,SRX26416452,SRS22936306,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,mate2-mate1 similar by mapping diff,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34016,SRR31031007,SRX26416451,SRS22936304,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,Zeb WT mDBF Rep3,GSM8578767,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:wildtype|treatment:minus DBF|geo loc name:missing|collection date:missing,Zeb WT mDBF Rep3,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:wildtype|treatment:minus DBF,GSM8578767,GSM8578767: Zeb WT mDBF Rep3; Danio rerio; OTHER,GSM8578767 r1,GSM8578767,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,Zeb_WT_mDBF_Rep3.R1.fq.gz Zeb_WT_mDBF_Rep3.R2.fq.gz,fastq fastq,20112719100.0,67042397.0,GSM8578767 r1,0:150 1:150,A:6537370232;C:2924919118;G:3973928235;T:6676228153;N:273362,150,150,,,6537370232,2924919118,3973928235,6676228153,273362,SRX26416451,SRS22936304,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,mate2-mate1 similar by mapping diff,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34017,SRR31031008,SRX26416450,SRS22936303,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,Zeb WT mDBF Rep2,GSM8578766,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:wildtype|treatment:minus DBF|geo loc name:missing|collection date:missing,Zeb WT mDBF Rep2,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:wildtype|treatment:minus DBF,GSM8578766,GSM8578766: Zeb WT mDBF Rep2; Danio rerio; OTHER,GSM8578766 r1,GSM8578766,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,Zeb_WT_mDBF_Rep2.R1.fq.gz Zeb_WT_mDBF_Rep2.R2.fq.gz,fastq fastq,24839552100.0,82798507.0,GSM8578766 r1,0:150 1:150,A:8176074473;C:3715794630;G:4984816273;T:7962528150;N:338574,150,150,,,8176074473,3715794630,4984816273,7962528150,338574,SRX26416450,SRS22936303,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,mate2-mate1 similar by mapping diff,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34018,SRR31031009,SRX26416449,SRS22936302,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,Zeb WT mDBF Rep1,GSM8578765,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:wildtype|treatment:minus DBF|geo loc name:missing|collection date:missing,Zeb WT mDBF Rep1,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:wildtype|treatment:minus DBF,GSM8578765,GSM8578765: Zeb WT mDBF Rep1; Danio rerio; OTHER,GSM8578765 r1,GSM8578765,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,Zeb_WT_mDBF_Rep1.R1.fq.gz Zeb_WT_mDBF_Rep1.R2.fq.gz,fastq fastq,23029785600.0,76765952.0,GSM8578765 r1,0:150 1:150,A:7509388471;C:3438475939;G:4820328116;T:7261277960;N:315114,150,150,,,7509388471,3438475939,4820328116,7261277960,315114,SRX26416449,SRS22936302,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,mate2-mate1 similar by mapping diff,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34019,SRR31031010,SRX26416448,SRS22936301,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,Zeb NES pDBF Rep2,GSM8578764,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF|geo loc name:missing|collection date:missing,Zeb NES pDBF Rep2,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF,GSM8578764,GSM8578764: Zeb NES pDBF Rep2; Danio rerio; OTHER,GSM8578764 r1,GSM8578764,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,Zeb_NES_pDBF_Rep2.R1.fq.gz Zeb_NES_pDBF_Rep2.R2.fq.gz,fastq fastq,25465584900.0,84885283.0,GSM8578764 r1,0:150 1:150,A:7944190976;C:4330095891;G:5473069331;T:7718120992;N:107710,150,150,,,7944190976,4330095891,5473069331,7718120992,107710,SRX26416448,SRS22936301,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,mate2-mate1 similar by mapping diff,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34020,SRR31031011,SRX26416447,SRS22936300,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,Zeb NES pDBF Rep1,GSM8578763,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF|geo loc name:missing|collection date:missing,Zeb NES pDBF Rep1,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF,GSM8578763,GSM8578763: Zeb NES pDBF Rep1; Danio rerio; OTHER,GSM8578763 r1,GSM8578763,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,Zeb_NES_pDBF_Rep1.R1.fq.gz Zeb_NES_pDBF_Rep1.R2.fq.gz,fastq fastq,21107351400.0,70357838.0,GSM8578763 r1,0:150 1:150,A:6528579219;C:3620666961;G:4496642830;T:6461373643;N:88747,150,150,,,6528579219,3620666961,4496642830,6461373643,88747,SRX26416447,SRS22936300,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,mate2-mate1 similar by mapping diff,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34021,SRR31031012,SRX26416446,SRS22936299,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,Zeb NES mDBF Rep2,GSM8578762,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF|geo loc name:missing|collection date:missing,Zeb NES mDBF Rep2,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF,GSM8578762,GSM8578762: Zeb NES mDBF Rep2; Danio rerio; OTHER,GSM8578762 r1,GSM8578762,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,Zeb_NES_mDBF_Rep2.R1.fq.gz Zeb_NES_mDBF_Rep2.R2.fq.gz,fastq fastq,21542967300.0,71809891.0,GSM8578762 r1,0:150 1:150,A:6722613760;C:3654507931;G:4628076548;T:6537678084;N:90977,150,150,,,6722613760,3654507931,4628076548,6537678084,90977,SRX26416446,SRS22936299,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,mate2-mate1 similar by mapping diff,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34022,SRR31031013,SRX26416445,SRS22936297,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,Zeb NES mDBF Rep1,GSM8578761,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF|geo loc name:missing|collection date:missing,Zeb NES mDBF Rep1,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF,GSM8578761,GSM8578761: Zeb NES mDBF Rep1; Danio rerio; OTHER,GSM8578761 r1,GSM8578761,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,Zeb_NES_mDBF_Rep1.R1.fq.gz Zeb_NES_mDBF_Rep1.R2.fq.gz,fastq fastq,23038870800.0,76796236.0,GSM8578761 r1,0:150 1:150,A:7216059153;C:3815899397;G:4909028200;T:7097787584;N:96466,150,150,,,7216059153,3815899397,4909028200,7097787584,96466,SRX26416445,SRS22936297,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,mate2-mate1 similar by mapping diff,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34023,SRR31031014,SRX26416444,SRS22936298,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF NES MALAT1 Rep4 plus,GSM8578760,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF|geo loc name:missing|collection date:missing,ZF NES MALAT1 Rep4 plus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF,GSM8578760,GSM8578760: ZF NES MALAT1 Rep4 plus; Danio rerio; OTHER,GSM8578760 r1,GSM8578760,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_NES_MALAT1_Rep4_plus.R1.fq.gz ZF_NES_MALAT1_Rep4_plus.R2.fq.gz,fastq fastq,1091293200.0,3637644.0,GSM8578760 r1,0:150 1:150,A:265519561;C:288647650;G:279105515;T:258001510;N:18964,150,150,,,265519561,288647650,279105515,258001510,18964,SRX26416444,SRS22936298,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34024,SRR31031015,SRX26416443,SRS22936296,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF NES MALAT1 Rep4 minus,GSM8578759,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF|geo loc name:missing|collection date:missing,ZF NES MALAT1 Rep4 minus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF,GSM8578759,GSM8578759: ZF NES MALAT1 Rep4 minus; Danio rerio; OTHER,GSM8578759 r1,GSM8578759,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_NES_MALAT1_Rep4_minus.R1.fq.gz ZF_NES_MALAT1_Rep4_minus.R2.fq.gz,fastq fastq,1395963600.0,4653212.0,GSM8578759 r1,0:150 1:150,A:339654289;C:369433701;G:356957685;T:329893853;N:24072,150,150,,,339654289,369433701,356957685,329893853,24072,SRX26416443,SRS22936296,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34025,SRR31031016,SRX26416442,SRS22936295,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF NES MALAT1 Rep3 plus,GSM8578758,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF|geo loc name:missing|collection date:missing,ZF NES MALAT1 Rep3 plus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF,GSM8578758,GSM8578758: ZF NES MALAT1 Rep3 plus; Danio rerio; OTHER,GSM8578758 r1,GSM8578758,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_NES_MALAT1_Rep3_plus.R1.fq.gz ZF_NES_MALAT1_Rep3_plus.R2.fq.gz,fastq fastq,1556195100.0,5187317.0,GSM8578758 r1,0:150 1:150,A:378553535;C:411730281;G:398086882;T:367797563;N:26839,150,150,,,378553535,411730281,398086882,367797563,26839,SRX26416442,SRS22936295,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34026,SRR31031017,SRX26416441,SRS22936294,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF NES MALAT1 Rep3 minus,GSM8578757,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF|geo loc name:missing|collection date:missing,ZF NES MALAT1 Rep3 minus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF,GSM8578757,GSM8578757: ZF NES MALAT1 Rep3 minus; Danio rerio; OTHER,GSM8578757 r1,GSM8578757,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_NES_MALAT1_Rep3_minus.R1.fq.gz ZF_NES_MALAT1_Rep3_minus.R2.fq.gz,fastq fastq,1511328900.0,5037763.0,GSM8578757 r1,0:150 1:150,A:367639521;C:400036694;G:386523395;T:357103340;N:25950,150,150,,,367639521,400036694,386523395,357103340,25950,SRX26416441,SRS22936294,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34027,SRR31031018,SRX26416440,SRS22936293,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF NES MALAT1 Rep2 plus,GSM8578756,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF|geo loc name:missing|collection date:missing,ZF NES MALAT1 Rep2 plus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF,GSM8578756,GSM8578756: ZF NES MALAT1 Rep2 plus; Danio rerio; OTHER,GSM8578756 r1,GSM8578756,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_NES_MALAT1_Rep2_plus.R1.fq.gz ZF_NES_MALAT1_Rep2_plus.R2.fq.gz,fastq fastq,1451137800.0,4837126.0,GSM8578756 r1,0:150 1:150,A:353052445;C:383981771;G:371154128;T:342925275;N:24181,150,150,,,353052445,383981771,371154128,342925275,24181,SRX26416440,SRS22936293,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34028,SRR31031019,SRX26416439,SRS22936292,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF NES MALAT1 Rep2 minus,GSM8578755,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF|geo loc name:missing|collection date:missing,ZF NES MALAT1 Rep2 minus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF,GSM8578755,GSM8578755: ZF NES MALAT1 Rep2 minus; Danio rerio; OTHER,GSM8578755 r1,GSM8578755,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_NES_MALAT1_Rep2_minus.R1.fq.gz ZF_NES_MALAT1_Rep2_minus.R2.fq.gz,fastq fastq,1286489400.0,4288298.0,GSM8578755 r1,0:150 1:150,A:312986127;C:340482608;G:329001591;T:303997982;N:21092,150,150,,,312986127,340482608,329001591,303997982,21092,SRX26416439,SRS22936292,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34029,SRR31031020,SRX26416438,SRS22936291,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF NES MALAT1 Rep1 plus,GSM8578754,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF|geo loc name:missing|collection date:missing,ZF NES MALAT1 Rep1 plus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF,GSM8578754,GSM8578754: ZF NES MALAT1 Rep1 plus; Danio rerio; OTHER,GSM8578754 r1,GSM8578754,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_NES_MALAT1_Rep1_plus.R1.fq.gz ZF_NES_MALAT1_Rep1_plus.R2.fq.gz,fastq fastq,1146380100.0,3821267.0,GSM8578754 r1,0:150 1:150,A:278826422;C:303464534;G:293239572;T:270830265;N:19307,150,150,,,278826422,303464534,293239572,270830265,19307,SRX26416438,SRS22936291,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34030,SRR31031021,SRX26416437,SRS22936289,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF NES MALAT1 Rep1 minus,GSM8578753,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF|geo loc name:missing|collection date:missing,ZF NES MALAT1 Rep1 minus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:minus DBF,GSM8578753,GSM8578753: ZF NES MALAT1 Rep1 minus; Danio rerio; OTHER,GSM8578753 r1,GSM8578753,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_NES_MALAT1_Rep1_minus.R1.fq.gz ZF_NES_MALAT1_Rep1_minus.R2.fq.gz,fastq fastq,1149087000.0,3830290.0,GSM8578753 r1,0:150 1:150,A:279487473;C:304130198;G:293947979;T:271501018;N:20332,150,150,,,279487473,304130198,293947979,271501018,20332,SRX26416437,SRS22936289,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
34031,SRR31031022,SRX26416436,SRS22936290,SRP539182,PRJNA1174121,Quantification of subcellular RNA localization through direct detection of RNA oxidation,GSE279714,Other,Across cell types and organisms  thousands of RNAs display asymmetric subcellular distributions. The study of this process often requires quantifying abundances of specific RNAs at precise subcellular locations. To analyze subcellular transcriptomes  multiple proximity based techniques have been developed in which RNAs near a localized bait protein are specifically labeled  facilitating their biotinylation and purification. However  these complex methods are often laborious and require expensive enrichment reagents. To streamline the analysis of localized RNA populations  we developed Oxidation Induced Nucleotide Conversion sequencing OINC seq. In OINC seq  RNAs near a genetically encoded  localized bait protein are specifically oxidized in a photo controllable manner. These oxidation events are then directly detected and quantified using high throughput sequencing and our software package  PIGPEN  without xxx need for biotin mediated enrichment. We demonstrate that OINC seq can induce and quantify RNA oxidation with high specificity in a dose  and light dependent manner. We further show the spatial specificity of OINC seq by using it to quantify subcellular transcriptomes associated with the cytoplasm  ER  and the inner and outer membranes of mitochondria. Finally  using transgenic zebrafish  we demonstrate that OINC seq allows proximity mediated RNA labeling in live animals. In sum  OINC seq together with PIGPEN provide an accessible workflow for the analysis of localized RNAs across different biological systems. Overall design: Cells and tissues were treated with and without xxx that drive localized RNA oxidation at a variety of subcellular locations  including treatment with the singlet oxygen producer Halo DBF and treatment with green light to activate Halo DBF. The extent of oxidation was then calculated at the gene level using PIGPEN software.,,pubmed:39605352;pubmed:40037712,,ZF NES GAPDH Rep4 plus,GSM8578752,,source name:2 dpf embryos|tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF|geo loc name:missing|collection date:missing,ZF NES GAPDH Rep4 plus,OINC seq Oxidation Induced Nucleotide Conversion sequencing Adapters were trimmed from reads using Cutadapt. RNA oxidation events as seen as mutations were quantified using PIGPEN https://github.com/TaliaferroLab/OINC seq Assembly: hg38 and GRCz11 Supplementary files format and content: Quantification of mutations in RNAseq data produced by PIGPEN.,2 dpf embryos,,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,HeLa cells were cultured in DMEM + pen/strep + 10% FBS. Zebrafish embryos were incubated at 28.5C in E3 medium.,tissue:2 dpf embryos|genotype:Halo NES|treatment:plus DBF,GSM8578752,GSM8578752: ZF NES GAPDH Rep4 plus; Danio rerio; OTHER,GSM8578752 r1,GSM8578752,1,RNA was extracted using Trizol and DNase treated for 30 min at 37C. Single amplicon libraries were constructed using gene specific RT PCR. Whole transcriptome libraries were created using Lexogen Quantseq library kits.,,OTHER,TRANSCRIPTOMIC,other,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP539182,,,ZF_NES_GAPDH_Rep4_plus.R1.fq.gz ZF_NES_GAPDH_Rep4_plus.R2.fq.gz,fastq fastq,1010253300.0,3367511.0,GSM8578752 r1,0:150 1:150,A:264284161;C:228880831;G:244038984;T:273032130;N:17194,150,150,,,264284161,228880831,244038984,273032130,17194,SRX26416436,SRS22936290,SRA1993027,"Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus","Biochemistry and Molecular Genetics, University of Colorado Anschutz Medical Campus",,,,,,,,,,,,B,B,biological fallback assumption,illumina,novaseq_era,full_length,random_priming,lexogen,bulk,unknown,unknown,,United States,2024-10-17,Hatching,Embryo,Embryo Imprecise,All anatomical structures
36409,SRR516560,SRX156356,SRS347213,SRP013950,PRJNA169500,Danio rerio embryonic promoterome,PRJNA169500,Transcriptome Analysis,Goal of this study is to generate genome wide maps of transcription initiation throughout early embryonic development of zebrafish Danio rerio. Cap analysis of gene expression CAGE is used to detect transcription start sites at 1bp resolution. CAGE data is complemented by ChIPseq datasets for promoter associated histone modifications to study dynamic changes of promoter usage and chromatin configuration throughout early embryonic development.,,pubmed:24531765,Zebrafish wild type AB strain   embryo prim20 stage,D. rerio prim20 embryo,D. rerio prim20 embryo,,,,,,,,,,,CAGE   D. rerio prim20 embryo,CAGE   D. rerio prim20 embryo run2,D. rerio prim20 embryo,1,,,OTHER,TRANSCRIPTOMIC,CAGE,SINGLE,ILLUMINA,Illumina Genome Analyzer IIx,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>27</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,SRP013950,,,CAGE_prim20_run1.fastq,fastq,76770666.0,2843358.0,CAGE   D. rerio prim20 embryo run1,0:27,A:19320047;C:17585086;G:22511175;T:17354358;N:0,27,,,,19320047,17585086,22511175,17354358,0,SRX156356,SRS347213,SRA055273,University of Bergen,ZEPROME consortium,1,0.56472,,0.10044,,0.77469,,0.74192,,27,,B,,usable mapping rate,illumina,early_illumina,unknown,cage,unknown,bulk,unknown,unknown,,Unknown,2012-06-28,Undetermined,Embryo,Embryo Imprecise,All anatomical structures
36410,SRR516561,SRX156356,SRS347213,SRP013950,PRJNA169500,Danio rerio embryonic promoterome,PRJNA169500,Transcriptome Analysis,Goal of this study is to generate genome wide maps of transcription initiation throughout early embryonic development of zebrafish Danio rerio. Cap analysis of gene expression CAGE is used to detect transcription start sites at 1bp resolution. CAGE data is complemented by ChIPseq datasets for promoter associated histone modifications to study dynamic changes of promoter usage and chromatin configuration throughout early embryonic development.,,pubmed:24531765,Zebrafish wild type AB strain   embryo prim20 stage,D. rerio prim20 embryo,D. rerio prim20 embryo,,,,,,,,,,,CAGE   D. rerio prim20 embryo,CAGE   D. rerio prim20 embryo run2,D. rerio prim20 embryo,1,,,OTHER,TRANSCRIPTOMIC,CAGE,SINGLE,ILLUMINA,Illumina Genome Analyzer IIx,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>27</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,SRP013950,,,CAGE_prim20_run2.fastq,fastq,163254906.0,6046478.0,CAGE   D. rerio prim20 embryo run2,0:27,A:40211042;C:36959797;G:46866669;T:39217398;N:0,27,,,,40211042,36959797,46866669,39217398,0,SRX156356,SRS347213,SRA055273,University of Bergen,ZEPROME consortium,1,0.47475,,0.06947,,0.78409,,0.78014,,27,,B,,usable mapping rate,illumina,early_illumina,unknown,cage,unknown,bulk,unknown,unknown,,Unknown,2013-08-31,Undetermined,Embryo,Embryo Imprecise,All anatomical structures
36411,SRR516559,SRX156355,SRS347212,SRP013950,PRJNA169500,Danio rerio embryonic promoterome,PRJNA169500,Transcriptome Analysis,Goal of this study is to generate genome wide maps of transcription initiation throughout early embryonic development of zebrafish Danio rerio. Cap analysis of gene expression CAGE is used to detect transcription start sites at 1bp resolution. CAGE data is complemented by ChIPseq datasets for promoter associated histone modifications to study dynamic changes of promoter usage and chromatin configuration throughout early embryonic development.,,pubmed:24531765,Zebrafish wild type AB strain   embryo prim6 stage,D. rerio prim6 embryo,D. rerio prim6 embryo,,,,,,,,,,,CAGE   D. rerio prim6 embryo,CAGE   D. rerio prim6 embryo,D. rerio prim6 embryo,1,,,OTHER,TRANSCRIPTOMIC,CAGE,SINGLE,ILLUMINA,Illumina Genome Analyzer IIx,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>27</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,SRP013950,,,CAGE_prim6.fastq,fastq,283414275.0,10496825.0,CAGE   D. rerio prim6 embryo,0:27,A:70974303;C:66202040;G:80064124;T:66173808;N:0,27,,,,70974303,66202040,80064124,66173808,0,SRX156355,SRS347212,SRA055273,University of Bergen,ZEPROME consortium,1,0.53395,,0.07806,,0.78255,,0.77575,,27,,B,,usable mapping rate,illumina,early_illumina,unknown,cage,unknown,bulk,unknown,unknown,,Unknown,2013-08-31,Undetermined,Embryo,Embryo Imprecise,All anatomical structures
36412,SRR516557,SRX156352,SRS347211,SRP013950,PRJNA169500,Danio rerio embryonic promoterome,PRJNA169500,Transcriptome Analysis,Goal of this study is to generate genome wide maps of transcription initiation throughout early embryonic development of zebrafish Danio rerio. Cap analysis of gene expression CAGE is used to detect transcription start sites at 1bp resolution. CAGE data is complemented by ChIPseq datasets for promoter associated histone modifications to study dynamic changes of promoter usage and chromatin configuration throughout early embryonic development.,,pubmed:24531765,Zebrafish wild type AB strain   embryo 14 somites stage,D. rerio 14 somites embryo,D. rerio 14 somites embryo,,,,,,,,,,,CAGE   D. rerio 14 somites embryo,CAGE   D. rerio 14 somites embryo run2,D. rerio 14 somites embryo,1,,,OTHER,TRANSCRIPTOMIC,CAGE,SINGLE,ILLUMINA,Illumina Genome Analyzer IIx,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>27</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,SRP013950,,,,,73689048.0,2729224.0,CAGE   D. rerio 14 somites embryo run1,0:27,A:19506796;C:16238588;G:21636330;T:16307334;N:0,27,,,,19506796,16238588,21636330,16307334,0,SRX156352,SRS347211,SRA055273,University of Bergen,ZEPROME consortium,1,0.63348,,0.09461,,0.76199,,0.68697,,27,,B,,usable mapping rate,illumina,early_illumina,unknown,cage,unknown,bulk,unknown,unknown,,Unknown,2012-06-28,Segmentation,Embryo,Embryo Imprecise,All anatomical structures
36413,SRR516558,SRX156352,SRS347211,SRP013950,PRJNA169500,Danio rerio embryonic promoterome,PRJNA169500,Transcriptome Analysis,Goal of this study is to generate genome wide maps of transcription initiation throughout early embryonic development of zebrafish Danio rerio. Cap analysis of gene expression CAGE is used to detect transcription start sites at 1bp resolution. CAGE data is complemented by ChIPseq datasets for promoter associated histone modifications to study dynamic changes of promoter usage and chromatin configuration throughout early embryonic development.,,pubmed:24531765,Zebrafish wild type AB strain   embryo 14 somites stage,D. rerio 14 somites embryo,D. rerio 14 somites embryo,,,,,,,,,,,CAGE   D. rerio 14 somites embryo,CAGE   D. rerio 14 somites embryo run2,D. rerio 14 somites embryo,1,,,OTHER,TRANSCRIPTOMIC,CAGE,SINGLE,ILLUMINA,Illumina Genome Analyzer IIx,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>27</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,SRP013950,,,CAGE_14somites_run2.fastq,fastq,212381811.0,7865993.0,CAGE   D. rerio 14 somites embryo run2,0:27,A:53780820;C:47467464;G:59893810;T:51239717;N:0,27,,,,53780820,47467464,59893810,51239717,0,SRX156352,SRS347211,SRA055273,University of Bergen,ZEPROME consortium,1,0.5278,,0.0806,,0.77589,,0.72881,,27,,B,,usable mapping rate,illumina,early_illumina,unknown,cage,unknown,bulk,unknown,unknown,,Unknown,2012-06-28,Segmentation,Embryo,Embryo Imprecise,All anatomical structures
36414,SRR516555,SRX156350,SRS347210,SRP013950,PRJNA169500,Danio rerio embryonic promoterome,PRJNA169500,Transcriptome Analysis,Goal of this study is to generate genome wide maps of transcription initiation throughout early embryonic development of zebrafish Danio rerio. Cap analysis of gene expression CAGE is used to detect transcription start sites at 1bp resolution. CAGE data is complemented by ChIPseq datasets for promoter associated histone modifications to study dynamic changes of promoter usage and chromatin configuration throughout early embryonic development.,,pubmed:24531765,Zebrafish wild type AB strain   embryo shield stage,D. rerio shield embryo,D. rerio shield embryo,,,,,,,,,,,CAGE   D. rerio shield embryo,CAGE   D. rerio shield embryo run2,D. rerio shield embryo,1,,,OTHER,TRANSCRIPTOMIC,CAGE,SINGLE,ILLUMINA,Illumina Genome Analyzer IIx,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>27</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,SRP013950,,,,,81086886.0,3003218.0,CAGE   D. rerio shield embryo run1,0:27,A:20800196;C:18582826;G:23491216;T:18212648;N:0,27,,,,20800196,18582826,23491216,18212648,0,SRX156350,SRS347210,SRA055273,University of Bergen,ZEPROME consortium,1,0.60434,,0.09948,,0.80432,,0.72631,,27,,B,,usable mapping rate,illumina,early_illumina,unknown,cage,unknown,bulk,unknown,unknown,,Unknown,2012-06-28,Gastrula,Embryo,Embryo Imprecise,All anatomical structures
36415,SRR516556,SRX156350,SRS347210,SRP013950,PRJNA169500,Danio rerio embryonic promoterome,PRJNA169500,Transcriptome Analysis,Goal of this study is to generate genome wide maps of transcription initiation throughout early embryonic development of zebrafish Danio rerio. Cap analysis of gene expression CAGE is used to detect transcription start sites at 1bp resolution. CAGE data is complemented by ChIPseq datasets for promoter associated histone modifications to study dynamic changes of promoter usage and chromatin configuration throughout early embryonic development.,,pubmed:24531765,Zebrafish wild type AB strain   embryo shield stage,D. rerio shield embryo,D. rerio shield embryo,,,,,,,,,,,CAGE   D. rerio shield embryo,CAGE   D. rerio shield embryo run2,D. rerio shield embryo,1,,,OTHER,TRANSCRIPTOMIC,CAGE,SINGLE,ILLUMINA,Illumina Genome Analyzer IIx,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>27</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,SRP013950,,,CAGE_shield_run2.fastq,fastq,81675891.0,3025033.0,CAGE   D. rerio shield embryo run2,0:27,A:21590166;C:17662459;G:22692127;T:19731139;N:0,27,,,,21590166,17662459,22692127,19731139,0,SRX156350,SRS347210,SRA055273,University of Bergen,ZEPROME consortium,1,0.52624,,0.08062,,0.81793,,0.76291,,27,,B,,usable mapping rate,illumina,early_illumina,unknown,cage,unknown,bulk,unknown,unknown,,Unknown,2012-06-28,Gastrula,Embryo,Embryo Imprecise,All anatomical structures
36416,SRR516554,SRX156349,SRS347209,SRP013950,PRJNA169500,Danio rerio embryonic promoterome,PRJNA169500,Transcriptome Analysis,Goal of this study is to generate genome wide maps of transcription initiation throughout early embryonic development of zebrafish Danio rerio. Cap analysis of gene expression CAGE is used to detect transcription start sites at 1bp resolution. CAGE data is complemented by ChIPseq datasets for promoter associated histone modifications to study dynamic changes of promoter usage and chromatin configuration throughout early embryonic development.,,pubmed:24531765,Zebrafish wild type AB strain   embryo dome/zfs:0000015 stage,D. rerio dome/zfs:0000015 embryo,D. rerio dome/zfs:0000015 embryo,,,,,,,,,,,CAGE   D. rerio dome/zfs:0000015 embryo,CAGE   D. rerio dome/zfs:0000015 embryo,D. rerio dome/zfs:0000015 embryo,1,,,OTHER,TRANSCRIPTOMIC,CAGE,SINGLE,ILLUMINA,Illumina Genome Analyzer IIx,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>27</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,SRP013950,,,CAGE_30p_dome.fastq,fastq,165690819.0,6136697.0,CAGE   D. rerio dome/zfs:0000015 embryo,0:27,A:42721293;C:36492232;G:46154926;T:40322368;N:0,27,,,,42721293,36492232,46154926,40322368,0,SRX156349,SRS347209,SRA055273,University of Bergen,ZEPROME consortium,1,0.50236,,0.0905,,0.81824,,0.8156,,27,,B,,usable mapping rate,illumina,early_illumina,unknown,cage,unknown,bulk,unknown,unknown,,Unknown,2012-06-28,Blastula,Embryo,Embryo Imprecise,All anatomical structures
36417,SRR516552,SRX156347,SRS347208,SRP013950,PRJNA169500,Danio rerio embryonic promoterome,PRJNA169500,Transcriptome Analysis,Goal of this study is to generate genome wide maps of transcription initiation throughout early embryonic development of zebrafish Danio rerio. Cap analysis of gene expression CAGE is used to detect transcription start sites at 1bp resolution. CAGE data is complemented by ChIPseq datasets for promoter associated histone modifications to study dynamic changes of promoter usage and chromatin configuration throughout early embryonic development.,,pubmed:24531765,Zebrafish wild type AB strain   embryo sphere/dome stage,D. rerio sphere/dome embryo,D. rerio sphere/dome embryo,,,,,,,,,,,CAGE   D. rerio sphere/dome embryo,CAGE   D. rerio sphere/dome embryo run2,D. rerio sphere/dome embryo,1,,,OTHER,TRANSCRIPTOMIC,CAGE,SINGLE,ILLUMINA,Illumina Genome Analyzer IIx,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>27</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,SRP013950,,,CAGE_sphere_dome_run1.fastq,fastq,78937740.0,2923620.0,CAGE   D. rerio sphere/dome embryo run1,0:27,A:20705496;C:17020957;G:22237043;T:18974244;N:0,27,,,,20705496,17020957,22237043,18974244,0,SRX156347,SRS347208,SRA055273,University of Bergen,ZEPROME consortium,1,0.50917,,0.09193,,0.81523,,0.8084,,27,,B,,usable mapping rate,illumina,early_illumina,unknown,cage,unknown,bulk,unknown,unknown,,Unknown,2012-06-28,Blastula,Embryo,Embryo Imprecise,All anatomical structures
36418,SRR516553,SRX156347,SRS347208,SRP013950,PRJNA169500,Danio rerio embryonic promoterome,PRJNA169500,Transcriptome Analysis,Goal of this study is to generate genome wide maps of transcription initiation throughout early embryonic development of zebrafish Danio rerio. Cap analysis of gene expression CAGE is used to detect transcription start sites at 1bp resolution. CAGE data is complemented by ChIPseq datasets for promoter associated histone modifications to study dynamic changes of promoter usage and chromatin configuration throughout early embryonic development.,,pubmed:24531765,Zebrafish wild type AB strain   embryo sphere/dome stage,D. rerio sphere/dome embryo,D. rerio sphere/dome embryo,,,,,,,,,,,CAGE   D. rerio sphere/dome embryo,CAGE   D. rerio sphere/dome embryo run2,D. rerio sphere/dome embryo,1,,,OTHER,TRANSCRIPTOMIC,CAGE,SINGLE,ILLUMINA,Illumina Genome Analyzer IIx,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>27</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,SRP013950,,,CAGE_sphere_dome_run2.fastq,fastq,82071009.0,3039667.0,CAGE   D. rerio sphere/dome embryo run2,0:27,A:20823133;C:17914934;G:22973465;T:20359477;N:0,27,,,,20823133,17914934,22973465,20359477,0,SRX156347,SRS347208,SRA055273,University of Bergen,ZEPROME consortium,1,0.47213,,0.07583,,0.82384,,0.8313,,27,,B,,usable mapping rate,illumina,early_illumina,unknown,cage,unknown,bulk,unknown,unknown,,Unknown,2012-06-28,Blastula,Embryo,Embryo Imprecise,All anatomical structures
36419,SRR516551,SRX156338,SRS347207,SRP013950,PRJNA169500,Danio rerio embryonic promoterome,PRJNA169500,Transcriptome Analysis,Goal of this study is to generate genome wide maps of transcription initiation throughout early embryonic development of zebrafish Danio rerio. Cap analysis of gene expression CAGE is used to detect transcription start sites at 1bp resolution. CAGE data is complemented by ChIPseq datasets for promoter associated histone modifications to study dynamic changes of promoter usage and chromatin configuration throughout early embryonic development.,,pubmed:24531765,Zebrafish wild type AB strain   embryo oblong stage,D. rerio oblong embryo,D. rerio oblong embryo,,,,,,,,,,,CAGE   D. rerio oblong embryo,CAGE   D. rerio oblong embryo,D. rerio oblong embryo,1,,,OTHER,TRANSCRIPTOMIC,CAGE,SINGLE,ILLUMINA,Illumina Genome Analyzer IIx,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>27</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,SRP013950,,,,,135892080.0,5033040.0,CAGE   D. rerio oblong embryo,0:27,A:34565898;C:30442218;G:38967845;T:31916119;N:0,27,,,,34565898,30442218,38967845,31916119,0,SRX156338,SRS347207,SRA055273,University of Bergen,ZEPROME consortium,1,0.56265,,0.09335,,0.80068,,0.70759,,27,,B,,usable mapping rate,illumina,early_illumina,unknown,cage,unknown,bulk,unknown,unknown,,Unknown,2012-06-28,Blastula,Embryo,Embryo Imprecise,All anatomical structures
36420,SRR516550,SRX156337,SRS347206,SRP013950,PRJNA169500,Danio rerio embryonic promoterome,PRJNA169500,Transcriptome Analysis,Goal of this study is to generate genome wide maps of transcription initiation throughout early embryonic development of zebrafish Danio rerio. Cap analysis of gene expression CAGE is used to detect transcription start sites at 1bp resolution. CAGE data is complemented by ChIPseq datasets for promoter associated histone modifications to study dynamic changes of promoter usage and chromatin configuration throughout early embryonic development.,,pubmed:24531765,Zebrafish wild type AB strain   embryo high stage,D. rerio high embryo,D. rerio high embryo,,,,,,,,,,,CAGE   D. rerio high embryo,CAGE   D. rerio high embryo,D. rerio high embryo,1,,,OTHER,TRANSCRIPTOMIC,CAGE,SINGLE,ILLUMINA,Illumina Genome Analyzer IIx,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>27</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,SRP013950,,,CAGE_high.fastq,fastq,128592846.0,4762698.0,CAGE   D. rerio high embryo,0:27,A:33693838;C:27629256;G:36692263;T:30577489;N:0,27,,,,33693838,27629256,36692263,30577489,0,SRX156337,SRS347206,SRA055273,University of Bergen,ZEPROME consortium,1,0.53282,,0.08323,,0.80854,,0.77395,,27,,B,,usable mapping rate,illumina,early_illumina,unknown,cage,unknown,bulk,unknown,unknown,,Unknown,2013-08-31,Undetermined,Embryo,Embryo Imprecise,All anatomical structures
36421,SRR516549,SRX156336,SRS347204,SRP013950,PRJNA169500,Danio rerio embryonic promoterome,PRJNA169500,Transcriptome Analysis,Goal of this study is to generate genome wide maps of transcription initiation throughout early embryonic development of zebrafish Danio rerio. Cap analysis of gene expression CAGE is used to detect transcription start sites at 1bp resolution. CAGE data is complemented by ChIPseq datasets for promoter associated histone modifications to study dynamic changes of promoter usage and chromatin configuration throughout early embryonic development.,,pubmed:24531765,Zebrafish wild type AB strain   embryo 512 cells stage,D. rerio 512 cells embryo,D. rerio 512 cells embyo,,,,,,,,,,,CAGE   D. rerio 512 cells embryo,CAGE   D. rerio 512 cells embryo,D. rerio 512 cells embryo,1,,,OTHER,TRANSCRIPTOMIC,CAGE,SINGLE,ILLUMINA,Illumina Genome Analyzer IIx,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>27</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,SRP013950,,,CAGE_512cells.fastq,fastq,150228810.0,5564030.0,CAGE   D. rerio 512 cells embryo,0:27,A:37826960;C:33838922;G:43403014;T:35159914;N:0,27,,,,37826960,33838922,43403014,35159914,0,SRX156336,SRS347204,SRA055273,University of Bergen,ZEPROME consortium,1,0.57925,,0.08753,,0.79933,,0.72579,,27,,B,,usable mapping rate,illumina,early_illumina,unknown,cage,unknown,bulk,unknown,unknown,,Unknown,2012-06-28,Blastula,Embryo,Embryo Imprecise,All anatomical structures
36422,SRR516548,SRX156334,SRS347202,SRP013950,PRJNA169500,Danio rerio embryonic promoterome,PRJNA169500,Transcriptome Analysis,Goal of this study is to generate genome wide maps of transcription initiation throughout early embryonic development of zebrafish Danio rerio. Cap analysis of gene expression CAGE is used to detect transcription start sites at 1bp resolution. CAGE data is complemented by ChIPseq datasets for promoter associated histone modifications to study dynamic changes of promoter usage and chromatin configuration throughout early embryonic development.,,pubmed:24531765,Zebrafish wild type AB strain   embryo 64 cells stage,D. rerio 64 cells embryo,D. rerio 64 cells embryo,,,,,,,,,,,CAGE   D. rerio 64 cells embryo,CAGE   D. rerio 64 cells embryo,D. rerio 64 cells embryo,1,,,OTHER,TRANSCRIPTOMIC,CAGE,SINGLE,ILLUMINA,Illumina Genome Analyzer IIx,<SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>27</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR>,SRP013950,,,CAGE_64cells.fastq,fastq,162784188.0,6029044.0,CAGE   D. rerio 64 cells embryo,0:27,A:41192860;C:36547869;G:46848833;T:38194626;N:0,27,,,,41192860,36547869,46848833,38194626,0,SRX156334,SRS347202,SRA055273,University of Bergen,ZEPROME consortium,,,,,,,,,,,,B,,usable mapping rate,illumina,early_illumina,unknown,cage,unknown,bulk,unknown,unknown,,Unknown,2013-08-31,Cleavage,Embryo,Embryo Imprecise,All anatomical structures
40249,SRR3038049,SRX1494244,SRS1217111,SRP067641,PRJNA306648,Dynamic RNA protein interactions underlie the zebrafish maternal to zygotic transition,GSE76212,Other,We adapted the iCLIP method to zebrafish embryos to investigate activities of Hnrnpa1 during the maternal to zygotic transition Overall design: 3 and 4 biological replicates from zebrafish embryos before preZGA and during ZGA zygotic genome activation  respectively  post UV crosslinking. Negative control samples include 3 IgG replicates of UV crosslinked samples for both developmental stages examined,,pubmed:28381614,,IgG iCLIP zf ZGA rep3,GSM1976593,,tissue:zebrafish embryo|developmental stage:ZGA 1K cell stage|antibody:anti IgG|barcode sequence:NNNTGGCNN,IgG iCLIP zf ZGA rep3,Basecalls performed using CASAVA version 1.4 For detail on data processing see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959 Genome build: Zv9 version 75 Supplementary files format and content: .bed files describing significant crosslink sites derived from unique contain G in the file name and multi mapped contain M in the file name reads for combined replicates for each individual developmental stages and negative controls.,zebrafish embryo,Zebrafish embryos at the desired developmental stages were irradiated with UV light 254 nm to in vivo crosslink RNA protein interactions.,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,Zebrafish embryos were raised from WT AB strain and maintained using standard conditions.,developmental stage:ZGA 1K cell stage|antibody:anti IgG|barcode sequence:NNNTGGCNN,GSM1976593,GSM1976593: IgG iCLIP zf ZGA rep3; Danio rerio; OTHER,GSM1976593,,1,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,GEO Accession:GSM1976593,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP067641,,,iCLIP_IgG_zebrafishembryo_wt_1K_IgGcontrol_Dr_NNNTGGCNN_20140613_L4333_4.fq.gz,fastq,82722124.0,1088449.0,GSM1976593 r1,0:76,A:21959870;C:19048567;G:22289272;T:19403630;N:20785,76,,,,21959870,19048567,22289272,19403630,20785,SRX1494244,SRS1217111,SRA320959,GEO,"Molecular Biophysics and Biochemistry, Yale University",1,0.08124,,0.02181,,0.98591,,0.59183,,76,,B,,usable mapping rate,illumina,hiseq_era,3prime,other,unknown,bulk,clip,iclip,,United States,2015-12-21,Blastula,Embryo,Embryo Imprecise,All anatomical structures
40250,SRR3038048,SRX1494243,SRS1217112,SRP067641,PRJNA306648,Dynamic RNA protein interactions underlie the zebrafish maternal to zygotic transition,GSE76212,Other,We adapted the iCLIP method to zebrafish embryos to investigate activities of Hnrnpa1 during the maternal to zygotic transition Overall design: 3 and 4 biological replicates from zebrafish embryos before preZGA and during ZGA zygotic genome activation  respectively  post UV crosslinking. Negative control samples include 3 IgG replicates of UV crosslinked samples for both developmental stages examined,,pubmed:28381614,,IgG iCLIP zf ZGA rep2,GSM1976592,,tissue:zebrafish embryo|developmental stage:ZGA 1K cell stage|antibody:anti IgG|barcode sequence:NNNCCACNN,IgG iCLIP zf ZGA rep2,Basecalls performed using CASAVA version 1.4 For detail on data processing see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959 Genome build: Zv9 version 75 Supplementary files format and content: .bed files describing significant crosslink sites derived from unique contain G in the file name and multi mapped contain M in the file name reads for combined replicates for each individual developmental stages and negative controls.,zebrafish embryo,Zebrafish embryos at the desired developmental stages were irradiated with UV light 254 nm to in vivo crosslink RNA protein interactions.,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,Zebrafish embryos were raised from WT AB strain and maintained using standard conditions.,developmental stage:ZGA 1K cell stage|antibody:anti IgG|barcode sequence:NNNCCACNN,GSM1976592,GSM1976592: IgG iCLIP zf ZGA rep2; Danio rerio; OTHER,GSM1976592,,1,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,GEO Accession:GSM1976592,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP067641,,,iCLIP_IgG_zebrafishembryo_wt_1K_IgGcontrol_Dr_NNNCCACNN_20140613_L4333_2.fq.gz,fastq,50195036.0,660461.0,GSM1976592 r1,0:76,A:14249581;C:12927029;G:12969451;T:10036400;N:12575,76,,,,14249581,12927029,12969451,10036400,12575,SRX1494243,SRS1217112,SRA320959,GEO,"Molecular Biophysics and Biochemistry, Yale University",1,0.25447,,0.03871,,0.94556,,0.7523,,76,,B,,usable mapping rate,illumina,hiseq_era,3prime,other,unknown,bulk,clip,iclip,,United States,2015-12-21,Blastula,Embryo,Embryo Imprecise,All anatomical structures
40251,SRR3038047,SRX1494242,SRS1217113,SRP067641,PRJNA306648,Dynamic RNA protein interactions underlie the zebrafish maternal to zygotic transition,GSE76212,Other,We adapted the iCLIP method to zebrafish embryos to investigate activities of Hnrnpa1 during the maternal to zygotic transition Overall design: 3 and 4 biological replicates from zebrafish embryos before preZGA and during ZGA zygotic genome activation  respectively  post UV crosslinking. Negative control samples include 3 IgG replicates of UV crosslinked samples for both developmental stages examined,,pubmed:28381614,,IgG iCLIP zf ZGA rep1,GSM1976591,,tissue:zebrafish embryo|developmental stage:ZGA 1K cell stage|antibody:anti IgG|barcode sequence:NNNGGCGNN,IgG iCLIP zf ZGA rep1,Basecalls performed using CASAVA version 1.4 For detail on data processing see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959 Genome build: Zv9 version 75 Supplementary files format and content: .bed files describing significant crosslink sites derived from unique contain G in the file name and multi mapped contain M in the file name reads for combined replicates for each individual developmental stages and negative controls.,zebrafish embryo,Zebrafish embryos at the desired developmental stages were irradiated with UV light 254 nm to in vivo crosslink RNA protein interactions.,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,Zebrafish embryos were raised from WT AB strain and maintained using standard conditions.,developmental stage:ZGA 1K cell stage|antibody:anti IgG|barcode sequence:NNNGGCGNN,GSM1976591,GSM1976591: IgG iCLIP zf ZGA rep1; Danio rerio; OTHER,GSM1976591,,1,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,GEO Accession:GSM1976591,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP067641,,,iCLIP_IgG_zebrafishembryo_wt_1K_IgGcontrol_Dr_NNNGGCGNN_20140613_L4333_5.fq.gz,fastq,453449136.0,5966436.0,GSM1976591 r1,0:76,A:124726528;C:101139098;G:135573575;T:91900507;N:109428,76,,,,124726528,101139098,135573575,91900507,109428,SRX1494242,SRS1217113,SRA320959,GEO,"Molecular Biophysics and Biochemistry, Yale University",1,0.51353,,0.10729,,0.88069,,0.7159,,76,,B,,usable mapping rate,illumina,hiseq_era,3prime,other,unknown,bulk,clip,iclip,,United States,2015-12-21,Blastula,Embryo,Embryo Imprecise,All anatomical structures
40252,SRR3038046,SRX1494241,SRS1217114,SRP067641,PRJNA306648,Dynamic RNA protein interactions underlie the zebrafish maternal to zygotic transition,GSE76212,Other,We adapted the iCLIP method to zebrafish embryos to investigate activities of Hnrnpa1 during the maternal to zygotic transition Overall design: 3 and 4 biological replicates from zebrafish embryos before preZGA and during ZGA zygotic genome activation  respectively  post UV crosslinking. Negative control samples include 3 IgG replicates of UV crosslinked samples for both developmental stages examined,,pubmed:28381614,,IgG iCLIP zf preZGA rep3,GSM1976590,,tissue:zebrafish embryo|developmental stage:preZGA 32 cell stage|antibody:anti IgG|barcode sequence:NNNTGGCNN,IgG iCLIP zf preZGA rep3,Basecalls performed using CASAVA version 1.4 For detail on data processing see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959 Genome build: Zv9 version 75 Supplementary files format and content: .bed files describing significant crosslink sites derived from unique contain G in the file name and multi mapped contain M in the file name reads for combined replicates for each individual developmental stages and negative controls.,zebrafish embryo,Zebrafish embryos at the desired developmental stages were irradiated with UV light 254 nm to in vivo crosslink RNA protein interactions.,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,Zebrafish embryos were raised from WT AB strain and maintained using standard conditions.,developmental stage:preZGA 32 cell stage|antibody:anti IgG|barcode sequence:NNNTGGCNN,GSM1976590,GSM1976590: IgG iCLIP zf preZGA rep3; Danio rerio; OTHER,GSM1976590,,1,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,GEO Accession:GSM1976590,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP067641,,,iCLIP_IgG_Drembryo_wt_IgGcontrol_Dr_NNNTGGCNN_20130812_hnRNPA1_4.fq.gz,fastq,600183932.0,7897157.0,GSM1976590 r1,0:76,A:173843237;C:131372689;G:170729621;T:124187689;N:50696,76,,,,173843237,131372689,170729621,124187689,50696,SRX1494241,SRS1217114,SRA320959,GEO,"Molecular Biophysics and Biochemistry, Yale University",1,0.14511,,0.03659,,0.9332,,0.651,,76,,B,,usable mapping rate,illumina,hiseq_era,3prime,other,unknown,bulk,clip,iclip,,United States,2015-12-21,Cleavage,Embryo,Embryo Imprecise,All anatomical structures
40253,SRR3038045,SRX1494240,SRS1217115,SRP067641,PRJNA306648,Dynamic RNA protein interactions underlie the zebrafish maternal to zygotic transition,GSE76212,Other,We adapted the iCLIP method to zebrafish embryos to investigate activities of Hnrnpa1 during the maternal to zygotic transition Overall design: 3 and 4 biological replicates from zebrafish embryos before preZGA and during ZGA zygotic genome activation  respectively  post UV crosslinking. Negative control samples include 3 IgG replicates of UV crosslinked samples for both developmental stages examined,,pubmed:28381614,,IgG iCLIP zf preZGA rep2,GSM1976589,,tissue:zebrafish embryo|developmental stage:preZGA 32 cell stage|antibody:anti IgG|barcode sequence:NNNCCGGNN,IgG iCLIP zf preZGA rep2,Basecalls performed using CASAVA version 1.4 For detail on data processing see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959 Genome build: Zv9 version 75 Supplementary files format and content: .bed files describing significant crosslink sites derived from unique contain G in the file name and multi mapped contain M in the file name reads for combined replicates for each individual developmental stages and negative controls.,zebrafish embryo,Zebrafish embryos at the desired developmental stages were irradiated with UV light 254 nm to in vivo crosslink RNA protein interactions.,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,Zebrafish embryos were raised from WT AB strain and maintained using standard conditions.,developmental stage:preZGA 32 cell stage|antibody:anti IgG|barcode sequence:NNNCCGGNN,GSM1976589,GSM1976589: IgG iCLIP zf preZGA rep2; Danio rerio; OTHER,GSM1976589,,1,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,GEO Accession:GSM1976589,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP067641,,,iCLIP_IgG_Drembryo_wt_IgGcontrol_Dr_NNNCCGGNN_20130812_hnRNPA1_5.fq.gz,fastq,612280168.0,8056318.0,GSM1976589 r1,0:76,A:175101988;C:144456623;G:172272609;T:120400014;N:48934,76,,,,175101988,144456623,172272609,120400014,48934,SRX1494240,SRS1217115,SRA320959,GEO,"Molecular Biophysics and Biochemistry, Yale University",1,0.05322,,0.01649,,0.97656,,0.67242,,76,,B,,usable mapping rate,illumina,hiseq_era,3prime,other,unknown,bulk,clip,iclip,,United States,2015-12-21,Cleavage,Embryo,Embryo Imprecise,All anatomical structures
40254,SRR3038044,SRX1494239,SRS1217116,SRP067641,PRJNA306648,Dynamic RNA protein interactions underlie the zebrafish maternal to zygotic transition,GSE76212,Other,We adapted the iCLIP method to zebrafish embryos to investigate activities of Hnrnpa1 during the maternal to zygotic transition Overall design: 3 and 4 biological replicates from zebrafish embryos before preZGA and during ZGA zygotic genome activation  respectively  post UV crosslinking. Negative control samples include 3 IgG replicates of UV crosslinked samples for both developmental stages examined,,pubmed:28381614,,IgG iCLIP zf preZGA rep1,GSM1976588,,tissue:zebrafish embryo|developmental stage:preZGA 32 cell stage|antibody:anti IgG|barcode sequence:NNNGGCGNN,IgG iCLIP zf preZGA rep1,Basecalls performed using CASAVA version 1.4 For detail on data processing see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959 Genome build: Zv9 version 75 Supplementary files format and content: .bed files describing significant crosslink sites derived from unique contain G in the file name and multi mapped contain M in the file name reads for combined replicates for each individual developmental stages and negative controls.,zebrafish embryo,Zebrafish embryos at the desired developmental stages were irradiated with UV light 254 nm to in vivo crosslink RNA protein interactions.,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,Zebrafish embryos were raised from WT AB strain and maintained using standard conditions.,developmental stage:preZGA 32 cell stage|antibody:anti IgG|barcode sequence:NNNGGCGNN,GSM1976588,GSM1976588: IgG iCLIP zf preZGA rep1; Danio rerio; OTHER,GSM1976588,,1,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,GEO Accession:GSM1976588,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP067641,,,iCLIP_IgG_Drembryo_wt_IgGcontrol_Dr_NNNGGCGNN_20130812_hnRNPA1_1.fq-3.gz,fastq,12806760.0,168510.0,GSM1976588 r1,0:76,A:3652525;C:2758781;G:3842061;T:2552042;N:1351,76,,,,3652525,2758781,3842061,2552042,1351,SRX1494239,SRS1217116,SRA320959,GEO,"Molecular Biophysics and Biochemistry, Yale University",1,0.02784,,0.01181,,0.99813,,0.68217,,76,,B,,usable mapping rate,illumina,hiseq_era,3prime,other,unknown,bulk,clip,iclip,,United States,2015-12-21,Cleavage,Embryo,Embryo Imprecise,All anatomical structures
40255,SRR3038043,SRX1494238,SRS1217117,SRP067641,PRJNA306648,Dynamic RNA protein interactions underlie the zebrafish maternal to zygotic transition,GSE76212,Other,We adapted the iCLIP method to zebrafish embryos to investigate activities of Hnrnpa1 during the maternal to zygotic transition Overall design: 3 and 4 biological replicates from zebrafish embryos before preZGA and during ZGA zygotic genome activation  respectively  post UV crosslinking. Negative control samples include 3 IgG replicates of UV crosslinked samples for both developmental stages examined,,pubmed:28381614,,hnRNP A1 iCLIP zf ZGA rep4,GSM1976587,,tissue:zebrafish embryo|developmental stage:ZGA 1K cell stage|antibody:anti hnRNP A1|barcode sequence:NNNGGTCNN,hnRNP A1 iCLIP zf ZGA rep4,Basecalls performed using CASAVA version 1.4 For detail on data processing see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959 Genome build: Zv9 version 75 Supplementary files format and content: .bed files describing significant crosslink sites derived from unique contain G in the file name and multi mapped contain M in the file name reads for combined replicates for each individual developmental stages and negative controls.,zebrafish embryo,Zebrafish embryos at the desired developmental stages were irradiated with UV light 254 nm to in vivo crosslink RNA protein interactions.,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,Zebrafish embryos were raised from WT AB strain and maintained using standard conditions.,developmental stage:ZGA 1K cell stage|antibody:anti hnRNP A1|barcode sequence:NNNGGTCNN,GSM1976587,GSM1976587: hnRNP A1 iCLIP zf ZGA rep4; Danio rerio; OTHER,GSM1976587,,1,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,GEO Accession:GSM1976587,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP067641,,,iCLIP_hnRNPA1_zebrafishembryo_wt_1K_hnRNPA1_Dr_NNNGGTCNN_20140613_L4333_6.fq.gz,fastq,2884007568.0,37947468.0,GSM1976587 r1,0:76,A:936463452;C:557587142;G:746595510;T:642623181;N:738283,76,,,,936463452,557587142,746595510,642623181,738283,SRX1494238,SRS1217117,SRA320959,GEO,"Molecular Biophysics and Biochemistry, Yale University",1,0.38306,,0.08291,,0.80568,,0.62132,,76,,B,,usable mapping rate,illumina,hiseq_era,3prime,other,unknown,bulk,clip,iclip,,United States,2015-12-21,Blastula,Embryo,Embryo Imprecise,All anatomical structures
40256,SRR3038042,SRX1494237,SRS1217118,SRP067641,PRJNA306648,Dynamic RNA protein interactions underlie the zebrafish maternal to zygotic transition,GSE76212,Other,We adapted the iCLIP method to zebrafish embryos to investigate activities of Hnrnpa1 during the maternal to zygotic transition Overall design: 3 and 4 biological replicates from zebrafish embryos before preZGA and during ZGA zygotic genome activation  respectively  post UV crosslinking. Negative control samples include 3 IgG replicates of UV crosslinked samples for both developmental stages examined,,pubmed:28381614,,hnRNP A1 iCLIP zf ZGA rep3,GSM1976586,,tissue:zebrafish embryo|developmental stage:ZGA 1K cell stage|antibody:anti hnRNP A1|barcode sequence:NNNCAATNN,hnRNP A1 iCLIP zf ZGA rep3,Basecalls performed using CASAVA version 1.4 For detail on data processing see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959 Genome build: Zv9 version 75 Supplementary files format and content: .bed files describing significant crosslink sites derived from unique contain G in the file name and multi mapped contain M in the file name reads for combined replicates for each individual developmental stages and negative controls.,zebrafish embryo,Zebrafish embryos at the desired developmental stages were irradiated with UV light 254 nm to in vivo crosslink RNA protein interactions.,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,Zebrafish embryos were raised from WT AB strain and maintained using standard conditions.,developmental stage:ZGA 1K cell stage|antibody:anti hnRNP A1|barcode sequence:NNNCAATNN,GSM1976586,GSM1976586: hnRNP A1 iCLIP zf ZGA rep3; Danio rerio; OTHER,GSM1976586,,1,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,GEO Accession:GSM1976586,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP067641,,,iCLIP_hnRNPA1_zebrafishembryo_wt_1K_hnRNPA1_Dr_NNNCAATNN_20140613_L4333_3.fq.gz,fastq,3875071508.0,50987783.0,GSM1976586 r1,0:76,A:1328984506;C:775814536;G:956147632;T:813109146;N:1015688,76,,,,1328984506,775814536,956147632,813109146,1015688,SRX1494237,SRS1217118,SRA320959,GEO,"Molecular Biophysics and Biochemistry, Yale University",1,0.35787,,0.0904,,0.81874,,0.64706,,76,,B,,usable mapping rate,illumina,hiseq_era,3prime,other,unknown,bulk,clip,iclip,,United States,2015-12-21,Blastula,Embryo,Embryo Imprecise,All anatomical structures
40257,SRR3038041,SRX1494236,SRS1217119,SRP067641,PRJNA306648,Dynamic RNA protein interactions underlie the zebrafish maternal to zygotic transition,GSE76212,Other,We adapted the iCLIP method to zebrafish embryos to investigate activities of Hnrnpa1 during the maternal to zygotic transition Overall design: 3 and 4 biological replicates from zebrafish embryos before preZGA and during ZGA zygotic genome activation  respectively  post UV crosslinking. Negative control samples include 3 IgG replicates of UV crosslinked samples for both developmental stages examined,,pubmed:28381614,,hnRNP A1 iCLIP zf ZGA rep2,GSM1976585,,tissue:zebrafish embryo|developmental stage:ZGA 1K cell stage|antibody:anti hnRNP A1|barcode sequence:NNNTTGTNN,hnRNP A1 iCLIP zf ZGA rep2,Basecalls performed using CASAVA version 1.4 For detail on data processing see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959 Genome build: Zv9 version 75 Supplementary files format and content: .bed files describing significant crosslink sites derived from unique contain G in the file name and multi mapped contain M in the file name reads for combined replicates for each individual developmental stages and negative controls.,zebrafish embryo,Zebrafish embryos at the desired developmental stages were irradiated with UV light 254 nm to in vivo crosslink RNA protein interactions.,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,Zebrafish embryos were raised from WT AB strain and maintained using standard conditions.,developmental stage:ZGA 1K cell stage|antibody:anti hnRNP A1|barcode sequence:NNNTTGTNN,GSM1976585,GSM1976585: hnRNP A1 iCLIP zf ZGA rep2; Danio rerio; OTHER,GSM1976585,,1,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,GEO Accession:GSM1976585,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP067641,,,iCLIP_hnRNPA1_zebrafishembryo_wt_1K_hnRNPA1_Dr_NNNTTGTNN_20140613_L4333_1.fq.gz,fastq,3482039180.0,45816305.0,GSM1976585 r1,0:76,A:1093877493;C:647486562;G:914636686;T:825135975;N:902464,76,,,,1093877493,647486562,914636686,825135975,902464,SRX1494236,SRS1217119,SRA320959,GEO,"Molecular Biophysics and Biochemistry, Yale University",1,0.29908,,0.07356,,0.83459,,0.62976,,76,,B,,usable mapping rate,illumina,hiseq_era,3prime,other,unknown,bulk,clip,iclip,,United States,2015-12-21,Blastula,Embryo,Embryo Imprecise,All anatomical structures
40258,SRR3038040,SRX1494235,SRS1217120,SRP067641,PRJNA306648,Dynamic RNA protein interactions underlie the zebrafish maternal to zygotic transition,GSE76212,Other,We adapted the iCLIP method to zebrafish embryos to investigate activities of Hnrnpa1 during the maternal to zygotic transition Overall design: 3 and 4 biological replicates from zebrafish embryos before preZGA and during ZGA zygotic genome activation  respectively  post UV crosslinking. Negative control samples include 3 IgG replicates of UV crosslinked samples for both developmental stages examined,,pubmed:28381614,,hnRNP A1 iCLIP zf ZGA rep1,GSM1976584,,tissue:zebrafish embryo|developmental stage:ZGA 1K cell stage|antibody:anti hnRNP A1|barcode sequence:NNNGGTTNN,hnRNP A1 iCLIP zf ZGA rep1,Basecalls performed using CASAVA version 1.4 For detail on data processing see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959 Genome build: Zv9 version 75 Supplementary files format and content: .bed files describing significant crosslink sites derived from unique contain G in the file name and multi mapped contain M in the file name reads for combined replicates for each individual developmental stages and negative controls.,zebrafish embryo,Zebrafish embryos at the desired developmental stages were irradiated with UV light 254 nm to in vivo crosslink RNA protein interactions.,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,Zebrafish embryos were raised from WT AB strain and maintained using standard conditions.,developmental stage:ZGA 1K cell stage|antibody:anti hnRNP A1|barcode sequence:NNNGGTTNN,GSM1976584,GSM1976584: hnRNP A1 iCLIP zf ZGA rep1; Danio rerio; OTHER,GSM1976584,,1,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,GEO Accession:GSM1976584,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP067641,,,iCLIP_hnRNPA1_zebrafishembryo_wt_1K_hnRNPA1_Dr_NNNGGTTNN_20140613_L4333_7.fq.gz,fastq,3547711920.0,46680420.0,GSM1976584 r1,0:76,A:1105542969;C:658172853;G:958654426;T:824422930;N:918742,76,,,,1105542969,658172853,958654426,824422930,918742,SRX1494235,SRS1217120,SRA320959,GEO,"Molecular Biophysics and Biochemistry, Yale University",1,0.34257,,0.08694,,0.83055,,0.64965,,76,,B,,usable mapping rate,illumina,hiseq_era,3prime,other,unknown,bulk,clip,iclip,,United States,2015-12-21,Blastula,Embryo,Embryo Imprecise,All anatomical structures
40259,SRR3038039,SRX1494234,SRS1217121,SRP067641,PRJNA306648,Dynamic RNA protein interactions underlie the zebrafish maternal to zygotic transition,GSE76212,Other,We adapted the iCLIP method to zebrafish embryos to investigate activities of Hnrnpa1 during the maternal to zygotic transition Overall design: 3 and 4 biological replicates from zebrafish embryos before preZGA and during ZGA zygotic genome activation  respectively  post UV crosslinking. Negative control samples include 3 IgG replicates of UV crosslinked samples for both developmental stages examined,,pubmed:28381614,,hnRNP A1 iCLIP zf preZGA rep3,GSM1976583,,tissue:zebrafish embryo|developmental stage:preZGA 32 cell stage|antibody:anti hnRNP A1|barcode sequence:NNNCAATNN,hnRNP A1 iCLIP zf preZGA rep3,Basecalls performed using CASAVA version 1.4 For detail on data processing see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959 Genome build: Zv9 version 75 Supplementary files format and content: .bed files describing significant crosslink sites derived from unique contain G in the file name and multi mapped contain M in the file name reads for combined replicates for each individual developmental stages and negative controls.,zebrafish embryo,Zebrafish embryos at the desired developmental stages were irradiated with UV light 254 nm to in vivo crosslink RNA protein interactions.,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,Zebrafish embryos were raised from WT AB strain and maintained using standard conditions.,developmental stage:preZGA 32 cell stage|antibody:anti hnRNP A1|barcode sequence:NNNCAATNN,GSM1976583,GSM1976583: hnRNP A1 iCLIP zf preZGA rep3; Danio rerio; OTHER,GSM1976583,,1,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,GEO Accession:GSM1976583,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP067641,,,iCLIP_hnRNPA1_Drembryo_wt_hnRNPA1_Dr_NNNCAATNN_20130812_hnRNPA1_3.fq.gz,fastq,1944912200.0,25590950.0,GSM1976583 r1,0:76,A:611172831;C:390755201;G:517844203;T:424973726;N:166239,76,,,,611172831,390755201,517844203,424973726,166239,SRX1494234,SRS1217121,SRA320959,GEO,"Molecular Biophysics and Biochemistry, Yale University",1,0.23819,,0.06019,,0.85318,,0.72811,,76,,B,,usable mapping rate,illumina,hiseq_era,3prime,other,unknown,bulk,clip,iclip,,United States,2015-12-21,Cleavage,Embryo,Embryo Imprecise,All anatomical structures
40260,SRR3038038,SRX1494233,SRS1217122,SRP067641,PRJNA306648,Dynamic RNA protein interactions underlie the zebrafish maternal to zygotic transition,GSE76212,Other,We adapted the iCLIP method to zebrafish embryos to investigate activities of Hnrnpa1 during the maternal to zygotic transition Overall design: 3 and 4 biological replicates from zebrafish embryos before preZGA and during ZGA zygotic genome activation  respectively  post UV crosslinking. Negative control samples include 3 IgG replicates of UV crosslinked samples for both developmental stages examined,,pubmed:28381614,,hnRNP A1 iCLIP zf preZGA rep2,GSM1976582,,tissue:zebrafish embryo|developmental stage:preZGA 32 cell stage|antibody:anti hnRNP A1|barcode sequence:NNNTTGTNN,hnRNP A1 iCLIP zf preZGA rep2,Basecalls performed using CASAVA version 1.4 For detail on data processing see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959 Genome build: Zv9 version 75 Supplementary files format and content: .bed files describing significant crosslink sites derived from unique contain G in the file name and multi mapped contain M in the file name reads for combined replicates for each individual developmental stages and negative controls.,zebrafish embryo,Zebrafish embryos at the desired developmental stages were irradiated with UV light 254 nm to in vivo crosslink RNA protein interactions.,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,Zebrafish embryos were raised from WT AB strain and maintained using standard conditions.,developmental stage:preZGA 32 cell stage|antibody:anti hnRNP A1|barcode sequence:NNNTTGTNN,GSM1976582,GSM1976582: hnRNP A1 iCLIP zf preZGA rep2; Danio rerio; OTHER,GSM1976582,,1,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,GEO Accession:GSM1976582,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP067641,,,iCLIP_hnRNPA1_Drembryo_wt_hnRNPA1_Dr_NNNTTGTNN_20130812_hnRNPA1_6.fq.gz,fastq,1804236124.0,23739949.0,GSM1976582 r1,0:76,A:520750722;C:337743522;G:499109535;T:446481275;N:151070,76,,,,520750722,337743522,499109535,446481275,151070,SRX1494233,SRS1217122,SRA320959,GEO,"Molecular Biophysics and Biochemistry, Yale University",1,0.25184,,0.06217,,0.85687,,0.67912,,76,,B,,usable mapping rate,illumina,hiseq_era,3prime,other,unknown,bulk,clip,iclip,,United States,2015-12-21,Cleavage,Embryo,Embryo Imprecise,All anatomical structures
40261,SRR3038037,SRX1494232,SRS1217123,SRP067641,PRJNA306648,Dynamic RNA protein interactions underlie the zebrafish maternal to zygotic transition,GSE76212,Other,We adapted the iCLIP method to zebrafish embryos to investigate activities of Hnrnpa1 during the maternal to zygotic transition Overall design: 3 and 4 biological replicates from zebrafish embryos before preZGA and during ZGA zygotic genome activation  respectively  post UV crosslinking. Negative control samples include 3 IgG replicates of UV crosslinked samples for both developmental stages examined,,pubmed:28381614,,hnRNP A1 iCLIP zf preZGA rep1,GSM1976581,,tissue:zebrafish embryo|developmental stage:preZGA 32 cell stage|antibody:anti hnRNP A1|barcode sequence:NNNGGTTNN,hnRNP A1 iCLIP zf preZGA rep1,Basecalls performed using CASAVA version 1.4 For detail on data processing see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959 Genome build: Zv9 version 75 Supplementary files format and content: .bed files describing significant crosslink sites derived from unique contain G in the file name and multi mapped contain M in the file name reads for combined replicates for each individual developmental stages and negative controls.,zebrafish embryo,Zebrafish embryos at the desired developmental stages were irradiated with UV light 254 nm to in vivo crosslink RNA protein interactions.,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,Zebrafish embryos were raised from WT AB strain and maintained using standard conditions.,developmental stage:preZGA 32 cell stage|antibody:anti hnRNP A1|barcode sequence:NNNGGTTNN,GSM1976581,GSM1976581: hnRNP A1 iCLIP zf preZGA rep1; Danio rerio; OTHER,GSM1976581,,1,Zebrafish embryos were lysed and lysates treated with RNaseI to obtain optimal length of RNA fragments for cDNA library preparation. hnRNP A1 RNA complexes were immunopurified using mouse anti hnRNP A1 antibody and protein G dynabeads Invitrogen. Anti IgG antibody Sigma was used for negative control immunopurifications. post RNA dephosphorylation  three prime end linker ligation was performed. Protein RNA complexes were resolved on SDS PAGE and transferred to the nitrocellulose membrane. Region of the membrane above the predicted size of hnRNP A1 was cut and RNA extracted using proteise K treatment. Subsequently  RNA was reverse transcribed using RT primers harboring a barcode. cDNA was separated into three size selected fractions using 6% TBE Urea gel Life Technologies and circularized by ssDNA circligase. Specific oligo was annealed for BamHI restriction site formation. cDNA was linearized by BamHI digestion. Final cDNA libraries were generated by PCR amplification.. For more details see: König et al. 2010: iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution. Nat Struct Mol Biol PMID:20601959,GEO Accession:GSM1976581,OTHER,TRANSCRIPTOMIC,other,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP067641,,,iCLIP_hnRNPA1_Drembryo_wt_hnRNPA1_Dr_NNNGGTTNN_20130812_hnRNPA1_2.fq.gz,fastq,2155200476.0,28357901.0,GSM1976581 r1,0:76,A:646739355;C:394864638;G:606938658;T:506474548;N:183277,76,,,,646739355,394864638,606938658,506474548,183277,SRX1494232,SRS1217123,SRA320959,GEO,"Molecular Biophysics and Biochemistry, Yale University",1,0.24207,,0.05859,,0.84747,,0.70564,,76,,B,,usable mapping rate,illumina,hiseq_era,3prime,other,unknown,bulk,clip,iclip,,United States,2015-12-21,Cleavage,Embryo,Embryo Imprecise,All anatomical structures
43058,SRR8354695,SRX5165478,SRS4174524,SRP114782,PRJNA397065,mRNA structure dynamics identifies RNA remodelers and functional elements during embryogenesis,PRJNA397065,Other,RNA folding plays a crucial role in RNA function. However  our knowledge of the global structure of the transcriptome is limited to steady state conditions  hindering our understanding of how RNA structure dynamics influences gene function. Here  we have characterized mRNA structure dynamics during the maternal to zygotic transition in zebrafish. We observe that on a global level  translation guides structure rather than structure guides translation. We detect a decrease in structure in translated regions  and identify the ribosome as a major remodeler of RNA structure in vivo. In contrast  we find that three prime UTRs form highly folded structures in vivo  which can affect gene expression by modulating miRNA activity. Furthermore  we find that dynamic three prime UTR structures are enriched in RNA decay elements  including regulatory elements in nanog  and cyclin A1  key maternal factors orchestrating the maternal to zygotic transition. These results reveal a central role of RNA structure dynamics in gene regulatory programs during embryogenesis.,,,,DMS seq   RNA structure positive controls in vitro replicate 1,DMS seq   RNA positives ctrl in vitro B1 AG01489,,strain:TU/AB|age:3.7|dev stage:oblong|sex:pooled male and female|tissue:embryo|treatment:0.5% DMS for 10 min in vitro|molecule:RNA|condition:invitro DMS|replicate group:1|replicate:1|barcode:TCTC|BioSampleModel:Model organism or animal,,,,,,,,,DMS seq   RNA structure positive controls in vitro replicate 1,AG01489.1,AG01489.1,RNA,,,OTHER,TRANSCRIPTOMIC,unspecified,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP114782,,,AG01489.1_R1.fastq.gz,fastq,75703530.0,2718404.0,AG01489.1 R1.fastq.gz,0:27.85 1:0,A:18206335;C:19464567;G:21319876;T:16712584;N:168,27,0,,,18206335,19464567,21319876,16712584,168,SRX5165478,SRS4174524,SRA825010,Yale_Giraldez|Genetics,Yale_Giraldez_Group,1,0.02389,,0.00591,,0.97654,,0.4758,,25,,B,,usable mapping rate,illumina,hiseq_era,unknown,small_rna,unknown,bulk,unknown,unknown,,United States,2018-12-19,Blastula,Embryo,Embryo Imprecise,All anatomical structures
43059,SRR8354696,SRX5165477,SRS4174523,SRP114782,PRJNA397065,mRNA structure dynamics identifies RNA remodelers and functional elements during embryogenesis,PRJNA397065,Other,RNA folding plays a crucial role in RNA function. However  our knowledge of the global structure of the transcriptome is limited to steady state conditions  hindering our understanding of how RNA structure dynamics influences gene function. Here  we have characterized mRNA structure dynamics during the maternal to zygotic transition in zebrafish. We observe that on a global level  translation guides structure rather than structure guides translation. We detect a decrease in structure in translated regions  and identify the ribosome as a major remodeler of RNA structure in vivo. In contrast  we find that three prime UTRs form highly folded structures in vivo  which can affect gene expression by modulating miRNA activity. Furthermore  we find that dynamic three prime UTR structures are enriched in RNA decay elements  including regulatory elements in nanog  and cyclin A1  key maternal factors orchestrating the maternal to zygotic transition. These results reveal a central role of RNA structure dynamics in gene regulatory programs during embryogenesis.,,,,DMS seq   RNA structure positive controls in vitro replicate 2,DMS seq   RNA positives ctrl in vitro B2 AG01490,,strain:TU/AB|age:3.7|dev stage:oblong|sex:pooled male and female|tissue:embryo|treatment:0.5% DMS for 10 min in vitro|molecule:RNA|condition:invitro DMS|replicate group:1|replicate:2|barcode:GTGT|BioSampleModel:Model organism or animal,,,,,,,,,DMS seq   RNA structure positive controls in vitro replicate 2,AG01490.1,AG01490.1,RNA,,,OTHER,TRANSCRIPTOMIC,unspecified,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP114782,,,AG01490.1_R1.fastq.gz,fastq,295051199.0,10869746.0,AG01490.1 R1.fastq.gz,0:27.14 1:0,A:71900468;C:76519029;G:81589099;T:65042019;N:584,27,0,,,71900468,76519029,81589099,65042019,584,SRX5165477,SRS4174523,SRA825010,Yale_Giraldez|Genetics,Yale_Giraldez_Group,1,0.02362,,0.00528,,0.98173,,0.35096,,19,,B,,usable mapping rate,illumina,hiseq_era,unknown,small_rna,unknown,bulk,unknown,unknown,,United States,2018-12-19,Blastula,Embryo,Embryo Imprecise,All anatomical structures
43060,SRR8354697,SRX5165476,SRS4174522,SRP114782,PRJNA397065,mRNA structure dynamics identifies RNA remodelers and functional elements during embryogenesis,PRJNA397065,Other,RNA folding plays a crucial role in RNA function. However  our knowledge of the global structure of the transcriptome is limited to steady state conditions  hindering our understanding of how RNA structure dynamics influences gene function. Here  we have characterized mRNA structure dynamics during the maternal to zygotic transition in zebrafish. We observe that on a global level  translation guides structure rather than structure guides translation. We detect a decrease in structure in translated regions  and identify the ribosome as a major remodeler of RNA structure in vivo. In contrast  we find that three prime UTRs form highly folded structures in vivo  which can affect gene expression by modulating miRNA activity. Furthermore  we find that dynamic three prime UTR structures are enriched in RNA decay elements  including regulatory elements in nanog  and cyclin A1  key maternal factors orchestrating the maternal to zygotic transition. These results reveal a central role of RNA structure dynamics in gene regulatory programs during embryogenesis.,,,,Raw multiplex: DMS seq   RNA structure positive controls in vitro replicate 1;DMS seq   RNA structure positive controls in vitro replicate 2,Raw multiplex: DMS seq   RNA positives ctrl in vitro B1 AG01489;DMS seq   RNA positives ctrl in vitro B2 AG01490,,strain:TU/AB|age:3.7|dev stage:oblong|sex:pooled male and female|tissue:embryo|treatment:0.5% DMS for 10 min in vitro|molecule:RNA|condition:invitro DMS|replicate group:1|replicate:1;2|barcode:TCTC;GTGT|BioSampleModel:Model organism or animal,,,,,,,,,Raw multiplex: DMS seq   RNA structure positive controls in vitro replicate 1;DMS seq   RNA structure positive controls in vitro replicate 2,AG01489.1;AG01490.1,AG01489.1;AG01490.1,RNA,,,OTHER,TRANSCRIPTOMIC,unspecified,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP114782,,,C7MYYANXX_JBDX127_009_R1.fastq.gz,fastq,1115150128.0,14673028.0,C7MYYANXX JBDX127 009 R1.fastq.gz,0:76,A:313482382;C:276309346;G:291633361;T:233659719;N:65320,76,,,,313482382,276309346,291633361,233659719,65320,SRX5165476,SRS4174522,SRA825010,Yale_Giraldez|Genetics,Yale_Giraldez_Group,1,0.00546,,0.00047,,0.98602,,0.51535,,76,,T,,under 1.2% mapping rate,illumina,hiseq_era,unknown,small_rna,unknown,bulk,unknown,unknown,,United States,2018-12-19,Blastula,Embryo,Embryo Imprecise,All anatomical structures
43061,SRR7264565,SRX4168732,SRS3380685,SRP114782,PRJNA397065,mRNA structure dynamics identifies RNA remodelers and functional elements during embryogenesis,PRJNA397065,Other,RNA folding plays a crucial role in RNA function. However  our knowledge of the global structure of the transcriptome is limited to steady state conditions  hindering our understanding of how RNA structure dynamics influences gene function. Here  we have characterized mRNA structure dynamics during the maternal to zygotic transition in zebrafish. We observe that on a global level  translation guides structure rather than structure guides translation. We detect a decrease in structure in translated regions  and identify the ribosome as a major remodeler of RNA structure in vivo. In contrast  we find that three prime UTRs form highly folded structures in vivo  which can affect gene expression by modulating miRNA activity. Furthermore  we find that dynamic three prime UTR structures are enriched in RNA decay elements  including regulatory elements in nanog  and cyclin A1  key maternal factors orchestrating the maternal to zygotic transition. These results reveal a central role of RNA structure dynamics in gene regulatory programs during embryogenesis.,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT shield in vivo;DMS Seq   WT sphere in vivo;unrelated,Raw multiplex: dmsseq AG01046;dmsseq AG01047;dmsseq AG01049;unrelated,,strain:TU/AB|age:2.0;6.0;4.0|dev stage:64c;shield;sphere|sex:pooled male and female|tissue:embryo|treatment:DMS in vivo|molecule:RNA|selection:pA|condition:DMS|replicate group:14;16;15|replicate:2|barcode:CACA;TCTC;AGAG;GTGT|BioSampleModel:Model organism or animal,,,,,,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT shield in vivo;DMS Seq   WT sphere in vivo;unrelated,AG01046.2;AG01047.2;AG01049.2;unrelated,AG01046.2;AG01047.2;AG01049.2;unrelated,RNA,,,OTHER,TRANSCRIPTOMIC,unspecified,SINGLE,ILLUMINA,Illumina HiSeq 2500,,SRP114782,,,HA22NADXX_JBBS71_021_R1.fastq.gz,fastq,493969676.0,6499601.0,HA22NADXX JBBS71 021 R1.fastq.gz,0:76,A:149523181;C:122615352;G:117180131;T:97949905;N:6701107,76,,,,149523181,122615352,117180131,97949905,6701107,SRX4168732,SRS3380685,SRA715414,Yale_Giraldez|Genetics,Yale_Giraldez_Group,1,0.0,,0.0,,1.0,,,,76,,T,,under 1.2% mapping rate,illumina,hiseq_era,unknown,small_rna,unknown,bulk,unknown,unknown,,United States,2018-06-05,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
43062,SRR7264566,SRX4168731,SRS3380685,SRP114782,PRJNA397065,mRNA structure dynamics identifies RNA remodelers and functional elements during embryogenesis,PRJNA397065,Other,RNA folding plays a crucial role in RNA function. However  our knowledge of the global structure of the transcriptome is limited to steady state conditions  hindering our understanding of how RNA structure dynamics influences gene function. Here  we have characterized mRNA structure dynamics during the maternal to zygotic transition in zebrafish. We observe that on a global level  translation guides structure rather than structure guides translation. We detect a decrease in structure in translated regions  and identify the ribosome as a major remodeler of RNA structure in vivo. In contrast  we find that three prime UTRs form highly folded structures in vivo  which can affect gene expression by modulating miRNA activity. Furthermore  we find that dynamic three prime UTR structures are enriched in RNA decay elements  including regulatory elements in nanog  and cyclin A1  key maternal factors orchestrating the maternal to zygotic transition. These results reveal a central role of RNA structure dynamics in gene regulatory programs during embryogenesis.,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT shield in vivo;DMS Seq   WT sphere in vivo;unrelated,Raw multiplex: dmsseq AG01046;dmsseq AG01047;dmsseq AG01049;unrelated,,strain:TU/AB|age:2.0;6.0;4.0|dev stage:64c;shield;sphere|sex:pooled male and female|tissue:embryo|treatment:DMS in vivo|molecule:RNA|selection:pA|condition:DMS|replicate group:14;16;15|replicate:2|barcode:CACA;TCTC;AGAG;GTGT|BioSampleModel:Model organism or animal,,,,,,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT shield in vivo;DMS Seq   WT sphere in vivo;unrelated,AG01046.1;AG01047.1;AG01049.1;unrelated,AG01046.1;AG01047.1;AG01049.1;unrelated,RNA,,,OTHER,TRANSCRIPTOMIC,unspecified,SINGLE,ILLUMINA,Illumina HiSeq 2500,,SRP114782,,,HA1YWADXX_JBBS71_021_R1.fastq.gz,fastq,1444702012.0,19009237.0,HA1YWADXX JBBS71 021 R1.fastq.gz,0:76,A:442549747;C:365473385;G:344546726;T:292070542;N:61612,76,,,,442549747,365473385,344546726,292070542,61612,SRX4168731,SRS3380685,SRA715414,Yale_Giraldez|Genetics,Yale_Giraldez_Group,1,0.0,,0.0,,1.0,,,,76,,T,,under 1.2% mapping rate,illumina,hiseq_era,unknown,small_rna,unknown,bulk,unknown,unknown,,United States,2018-06-05,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
43063,SRR7264567,SRX4168730,SRS3380690,SRP114782,PRJNA397065,mRNA structure dynamics identifies RNA remodelers and functional elements during embryogenesis,PRJNA397065,Other,RNA folding plays a crucial role in RNA function. However  our knowledge of the global structure of the transcriptome is limited to steady state conditions  hindering our understanding of how RNA structure dynamics influences gene function. Here  we have characterized mRNA structure dynamics during the maternal to zygotic transition in zebrafish. We observe that on a global level  translation guides structure rather than structure guides translation. We detect a decrease in structure in translated regions  and identify the ribosome as a major remodeler of RNA structure in vivo. In contrast  we find that three prime UTRs form highly folded structures in vivo  which can affect gene expression by modulating miRNA activity. Furthermore  we find that dynamic three prime UTR structures are enriched in RNA decay elements  including regulatory elements in nanog  and cyclin A1  key maternal factors orchestrating the maternal to zygotic transition. These results reveal a central role of RNA structure dynamics in gene regulatory programs during embryogenesis.,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT sphere in vivo;DMS Seq   WT shield in vivo;unrelated,Raw multiplex: dmsseq AG01042;dmsseq AG01043;dmsseq AG01044;unrelated,,strain:TU/AB|age:2.0;4.0;6.0|dev stage:64c;sphere;shield|sex:pooled male and female|tissue:embryo|treatment:DMS in vivo|molecule:RNA|selection:pA|condition:DMS|replicate group:14;15;16|replicate:1|barcode:CACA;AGAG;TCTC;GTGT|BioSampleModel:Model organism or animal,,,,,,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT sphere in vivo;DMS Seq   WT shield in vivo;unrelated,AG01042.6;AG01043.6;AG01044.6;unrelated,AG01042.6;AG01043.6;AG01044.6;unrelated,RNA,,,OTHER,TRANSCRIPTOMIC,unspecified,SINGLE,ILLUMINA,Illumina HiSeq 2500,,SRP114782,,,HBCAFADXX_JBBR70_020_R1.fastq.gz,fastq,3562176468.0,46870743.0,HBCAFADXX JBBR70 020 R1.fastq.gz,0:76,A:1097921080;C:906895047;G:849908073;T:707369464;N:82804,76,,,,1097921080,906895047,849908073,707369464,82804,SRX4168730,SRS3380690,SRA715414,Yale_Giraldez|Genetics,Yale_Giraldez_Group,1,0.0,,0.0,,1.0,,,,76,,T,,under 1.2% mapping rate,illumina,hiseq_era,unknown,small_rna,unknown,bulk,unknown,unknown,,United States,2018-06-05,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
43064,SRR7264568,SRX4168729,SRS3380690,SRP114782,PRJNA397065,mRNA structure dynamics identifies RNA remodelers and functional elements during embryogenesis,PRJNA397065,Other,RNA folding plays a crucial role in RNA function. However  our knowledge of the global structure of the transcriptome is limited to steady state conditions  hindering our understanding of how RNA structure dynamics influences gene function. Here  we have characterized mRNA structure dynamics during the maternal to zygotic transition in zebrafish. We observe that on a global level  translation guides structure rather than structure guides translation. We detect a decrease in structure in translated regions  and identify the ribosome as a major remodeler of RNA structure in vivo. In contrast  we find that three prime UTRs form highly folded structures in vivo  which can affect gene expression by modulating miRNA activity. Furthermore  we find that dynamic three prime UTR structures are enriched in RNA decay elements  including regulatory elements in nanog  and cyclin A1  key maternal factors orchestrating the maternal to zygotic transition. These results reveal a central role of RNA structure dynamics in gene regulatory programs during embryogenesis.,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT sphere in vivo;DMS Seq   WT shield in vivo;unrelated,Raw multiplex: dmsseq AG01042;dmsseq AG01043;dmsseq AG01044;unrelated,,strain:TU/AB|age:2.0;4.0;6.0|dev stage:64c;sphere;shield|sex:pooled male and female|tissue:embryo|treatment:DMS in vivo|molecule:RNA|selection:pA|condition:DMS|replicate group:14;15;16|replicate:1|barcode:CACA;AGAG;TCTC;GTGT|BioSampleModel:Model organism or animal,,,,,,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT sphere in vivo;DMS Seq   WT shield in vivo;unrelated,AG01042.5;AG01043.5;AG01044.5;unrelated,AG01042.5;AG01043.5;AG01044.5;unrelated,RNA,,,OTHER,TRANSCRIPTOMIC,unspecified,SINGLE,ILLUMINA,Illumina HiSeq 2500,,SRP114782,,,C562AACXX_JBBR70_NOBCX_R1.fastq.gz,fastq,13352386192.0,175689292.0,C562AACXX JBBR70 NOBCX R1.fastq.gz,0:76,A:4126007145;C:3372127203;G:3172182791;T:2661287884;N:20781169,76,,,,4126007145,3372127203,3172182791,2661287884,20781169,SRX4168729,SRS3380690,SRA715414,Yale_Giraldez|Genetics,Yale_Giraldez_Group,1,0.0,,0.0,,1.0,,,,76,,T,,under 1.2% mapping rate,illumina,hiseq_era,unknown,small_rna,unknown,bulk,unknown,unknown,,United States,2018-06-07,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
43065,SRR7264569,SRX4168728,SRS3380690,SRP114782,PRJNA397065,mRNA structure dynamics identifies RNA remodelers and functional elements during embryogenesis,PRJNA397065,Other,RNA folding plays a crucial role in RNA function. However  our knowledge of the global structure of the transcriptome is limited to steady state conditions  hindering our understanding of how RNA structure dynamics influences gene function. Here  we have characterized mRNA structure dynamics during the maternal to zygotic transition in zebrafish. We observe that on a global level  translation guides structure rather than structure guides translation. We detect a decrease in structure in translated regions  and identify the ribosome as a major remodeler of RNA structure in vivo. In contrast  we find that three prime UTRs form highly folded structures in vivo  which can affect gene expression by modulating miRNA activity. Furthermore  we find that dynamic three prime UTR structures are enriched in RNA decay elements  including regulatory elements in nanog  and cyclin A1  key maternal factors orchestrating the maternal to zygotic transition. These results reveal a central role of RNA structure dynamics in gene regulatory programs during embryogenesis.,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT sphere in vivo;DMS Seq   WT shield in vivo;unrelated,Raw multiplex: dmsseq AG01042;dmsseq AG01043;dmsseq AG01044;unrelated,,strain:TU/AB|age:2.0;4.0;6.0|dev stage:64c;sphere;shield|sex:pooled male and female|tissue:embryo|treatment:DMS in vivo|molecule:RNA|selection:pA|condition:DMS|replicate group:14;15;16|replicate:1|barcode:CACA;AGAG;TCTC;GTGT|BioSampleModel:Model organism or animal,,,,,,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT sphere in vivo;DMS Seq   WT shield in vivo;unrelated,AG01042.4;AG01043.4;AG01044.4;unrelated,AG01042.4;AG01043.4;AG01044.4;unrelated,RNA,,,OTHER,TRANSCRIPTOMIC,unspecified,SINGLE,ILLUMINA,Illumina HiSeq 2500,,SRP114782,,,HA2H1ADXX_JBBR70_NOBCX_R1.fastq.gz,fastq,9139389880.0,120255130.0,HA2H1ADXX JBBR70 NOBCX R1.fastq.gz,0:76,A:2815354570;C:2322972018;G:2180978488;T:1817457488;N:2627316,76,,,,2815354570,2322972018,2180978488,1817457488,2627316,SRX4168728,SRS3380690,SRA715414,Yale_Giraldez|Genetics,Yale_Giraldez_Group,1,0.0,,0.0,,1.0,,,,76,,T,,under 1.2% mapping rate,illumina,hiseq_era,unknown,small_rna,unknown,bulk,unknown,unknown,,United States,2018-06-07,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
43066,SRR7264570,SRX4168727,SRS3380690,SRP114782,PRJNA397065,mRNA structure dynamics identifies RNA remodelers and functional elements during embryogenesis,PRJNA397065,Other,RNA folding plays a crucial role in RNA function. However  our knowledge of the global structure of the transcriptome is limited to steady state conditions  hindering our understanding of how RNA structure dynamics influences gene function. Here  we have characterized mRNA structure dynamics during the maternal to zygotic transition in zebrafish. We observe that on a global level  translation guides structure rather than structure guides translation. We detect a decrease in structure in translated regions  and identify the ribosome as a major remodeler of RNA structure in vivo. In contrast  we find that three prime UTRs form highly folded structures in vivo  which can affect gene expression by modulating miRNA activity. Furthermore  we find that dynamic three prime UTR structures are enriched in RNA decay elements  including regulatory elements in nanog  and cyclin A1  key maternal factors orchestrating the maternal to zygotic transition. These results reveal a central role of RNA structure dynamics in gene regulatory programs during embryogenesis.,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT sphere in vivo;DMS Seq   WT shield in vivo;unrelated,Raw multiplex: dmsseq AG01042;dmsseq AG01043;dmsseq AG01044;unrelated,,strain:TU/AB|age:2.0;4.0;6.0|dev stage:64c;sphere;shield|sex:pooled male and female|tissue:embryo|treatment:DMS in vivo|molecule:RNA|selection:pA|condition:DMS|replicate group:14;15;16|replicate:1|barcode:CACA;AGAG;TCTC;GTGT|BioSampleModel:Model organism or animal,,,,,,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT sphere in vivo;DMS Seq   WT shield in vivo;unrelated,AG01042.3;AG01043.3;AG01044.3;unrelated,AG01042.3;AG01043.3;AG01044.3;unrelated,RNA,,,OTHER,TRANSCRIPTOMIC,unspecified,SINGLE,ILLUMINA,Illumina HiSeq 2500,,SRP114782,,,HA1YVADXX_JBBR70_NOBCX_R1.fastq.gz,fastq,7892000132.0,103842107.0,HA1YVADXX JBBR70 NOBCX R1.fastq.gz,0:76,A:2428266964;C:2007082407;G:1885147455;T:1568726692;N:2776614,76,,,,2428266964,2007082407,1885147455,1568726692,2776614,SRX4168727,SRS3380690,SRA715414,Yale_Giraldez|Genetics,Yale_Giraldez_Group,1,0.0,,0.0,,1.0,,,,76,,T,,under 1.2% mapping rate,illumina,hiseq_era,unknown,small_rna,unknown,bulk,unknown,unknown,,United States,2018-06-05,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
43067,SRR7264571,SRX4168726,SRS3380690,SRP114782,PRJNA397065,mRNA structure dynamics identifies RNA remodelers and functional elements during embryogenesis,PRJNA397065,Other,RNA folding plays a crucial role in RNA function. However  our knowledge of the global structure of the transcriptome is limited to steady state conditions  hindering our understanding of how RNA structure dynamics influences gene function. Here  we have characterized mRNA structure dynamics during the maternal to zygotic transition in zebrafish. We observe that on a global level  translation guides structure rather than structure guides translation. We detect a decrease in structure in translated regions  and identify the ribosome as a major remodeler of RNA structure in vivo. In contrast  we find that three prime UTRs form highly folded structures in vivo  which can affect gene expression by modulating miRNA activity. Furthermore  we find that dynamic three prime UTR structures are enriched in RNA decay elements  including regulatory elements in nanog  and cyclin A1  key maternal factors orchestrating the maternal to zygotic transition. These results reveal a central role of RNA structure dynamics in gene regulatory programs during embryogenesis.,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT sphere in vivo;DMS Seq   WT shield in vivo;unrelated,Raw multiplex: dmsseq AG01042;dmsseq AG01043;dmsseq AG01044;unrelated,,strain:TU/AB|age:2.0;4.0;6.0|dev stage:64c;sphere;shield|sex:pooled male and female|tissue:embryo|treatment:DMS in vivo|molecule:RNA|selection:pA|condition:DMS|replicate group:14;15;16|replicate:1|barcode:CACA;AGAG;TCTC;GTGT|BioSampleModel:Model organism or animal,,,,,,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT sphere in vivo;DMS Seq   WT shield in vivo;unrelated,AG01042.2;AG01043.2;AG01044.2;unrelated,AG01042.2;AG01043.2;AG01044.2;unrelated,RNA,,,OTHER,TRANSCRIPTOMIC,unspecified,SINGLE,ILLUMINA,Illumina HiSeq 2500,,SRP114782,,,HA22NADXX_JBBR70_020_R1.fastq.gz,fastq,554569568.0,7296968.0,HA22NADXX JBBR70 020 R1.fastq.gz,0:76,A:168630291;C:138451887;G:131779810;T:108186590;N:7520990,76,,,,168630291,138451887,131779810,108186590,7520990,SRX4168726,SRS3380690,SRA715414,Yale_Giraldez|Genetics,Yale_Giraldez_Group,1,0.0,,0.0,,1.0,,,,76,,T,,under 1.2% mapping rate,illumina,hiseq_era,unknown,small_rna,unknown,bulk,unknown,unknown,,United States,2018-06-05,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
43068,SRR7264572,SRX4168725,SRS3380690,SRP114782,PRJNA397065,mRNA structure dynamics identifies RNA remodelers and functional elements during embryogenesis,PRJNA397065,Other,RNA folding plays a crucial role in RNA function. However  our knowledge of the global structure of the transcriptome is limited to steady state conditions  hindering our understanding of how RNA structure dynamics influences gene function. Here  we have characterized mRNA structure dynamics during the maternal to zygotic transition in zebrafish. We observe that on a global level  translation guides structure rather than structure guides translation. We detect a decrease in structure in translated regions  and identify the ribosome as a major remodeler of RNA structure in vivo. In contrast  we find that three prime UTRs form highly folded structures in vivo  which can affect gene expression by modulating miRNA activity. Furthermore  we find that dynamic three prime UTR structures are enriched in RNA decay elements  including regulatory elements in nanog  and cyclin A1  key maternal factors orchestrating the maternal to zygotic transition. These results reveal a central role of RNA structure dynamics in gene regulatory programs during embryogenesis.,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT sphere in vivo;DMS Seq   WT shield in vivo;unrelated,Raw multiplex: dmsseq AG01042;dmsseq AG01043;dmsseq AG01044;unrelated,,strain:TU/AB|age:2.0;4.0;6.0|dev stage:64c;sphere;shield|sex:pooled male and female|tissue:embryo|treatment:DMS in vivo|molecule:RNA|selection:pA|condition:DMS|replicate group:14;15;16|replicate:1|barcode:CACA;AGAG;TCTC;GTGT|BioSampleModel:Model organism or animal,,,,,,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT sphere in vivo;DMS Seq   WT shield in vivo;unrelated,AG01042.1;AG01043.1;AG01044.1;unrelated,AG01042.1;AG01043.1;AG01044.1;unrelated,RNA,,,OTHER,TRANSCRIPTOMIC,unspecified,SINGLE,ILLUMINA,Illumina HiSeq 2500,,SRP114782,,,HA1YWADXX_JBBR70_020_R1.fastq.gz,fastq,1565371316.0,20596991.0,HA1YWADXX JBBR70 020 R1.fastq.gz,0:76,A:481887814;C:398331513;G:374271346;T:310815367;N:65276,76,,,,481887814,398331513,374271346,310815367,65276,SRX4168725,SRS3380690,SRA715414,Yale_Giraldez|Genetics,Yale_Giraldez_Group,1,0.0,,0.0,,1.0,,,,76,,T,,under 1.2% mapping rate,illumina,hiseq_era,unknown,small_rna,unknown,bulk,unknown,unknown,,United States,2018-06-05,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
43069,SRR7264573,SRX4168724,SRS3380685,SRP114782,PRJNA397065,mRNA structure dynamics identifies RNA remodelers and functional elements during embryogenesis,PRJNA397065,Other,RNA folding plays a crucial role in RNA function. However  our knowledge of the global structure of the transcriptome is limited to steady state conditions  hindering our understanding of how RNA structure dynamics influences gene function. Here  we have characterized mRNA structure dynamics during the maternal to zygotic transition in zebrafish. We observe that on a global level  translation guides structure rather than structure guides translation. We detect a decrease in structure in translated regions  and identify the ribosome as a major remodeler of RNA structure in vivo. In contrast  we find that three prime UTRs form highly folded structures in vivo  which can affect gene expression by modulating miRNA activity. Furthermore  we find that dynamic three prime UTR structures are enriched in RNA decay elements  including regulatory elements in nanog  and cyclin A1  key maternal factors orchestrating the maternal to zygotic transition. These results reveal a central role of RNA structure dynamics in gene regulatory programs during embryogenesis.,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT shield in vivo;DMS Seq   WT sphere in vivo;unrelated,Raw multiplex: dmsseq AG01046;dmsseq AG01047;dmsseq AG01049;unrelated,,strain:TU/AB|age:2.0;6.0;4.0|dev stage:64c;shield;sphere|sex:pooled male and female|tissue:embryo|treatment:DMS in vivo|molecule:RNA|selection:pA|condition:DMS|replicate group:14;16;15|replicate:2|barcode:CACA;TCTC;AGAG;GTGT|BioSampleModel:Model organism or animal,,,,,,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT shield in vivo;DMS Seq   WT sphere in vivo;unrelated,AG01046.4;AG01047.4;AG01049.4;unrelated,AG01046.4;AG01047.4;AG01049.4;unrelated,RNA,,,OTHER,TRANSCRIPTOMIC,unspecified,SINGLE,ILLUMINA,Illumina HiSeq 2500,,SRP114782,,,HA2H1ADXX_JBBS71_NOBCX_R1.fastq.gz,fastq,8845878184.0,116393134.0,HA2H1ADXX JBBS71 NOBCX R1.fastq.gz,0:76,A:2711637569;C:2235324807;G:2105535243;T:1791610098;N:1770467,76,,,,2711637569,2235324807,2105535243,1791610098,1770467,SRX4168724,SRS3380685,SRA715414,Yale_Giraldez|Genetics,Yale_Giraldez_Group,1,0.0,,0.0,,1.0,,,,76,,T,,under 1.2% mapping rate,illumina,hiseq_era,unknown,small_rna,unknown,bulk,unknown,unknown,,United States,2018-06-07,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
43070,SRR7264574,SRX4168723,SRS3380685,SRP114782,PRJNA397065,mRNA structure dynamics identifies RNA remodelers and functional elements during embryogenesis,PRJNA397065,Other,RNA folding plays a crucial role in RNA function. However  our knowledge of the global structure of the transcriptome is limited to steady state conditions  hindering our understanding of how RNA structure dynamics influences gene function. Here  we have characterized mRNA structure dynamics during the maternal to zygotic transition in zebrafish. We observe that on a global level  translation guides structure rather than structure guides translation. We detect a decrease in structure in translated regions  and identify the ribosome as a major remodeler of RNA structure in vivo. In contrast  we find that three prime UTRs form highly folded structures in vivo  which can affect gene expression by modulating miRNA activity. Furthermore  we find that dynamic three prime UTR structures are enriched in RNA decay elements  including regulatory elements in nanog  and cyclin A1  key maternal factors orchestrating the maternal to zygotic transition. These results reveal a central role of RNA structure dynamics in gene regulatory programs during embryogenesis.,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT shield in vivo;DMS Seq   WT sphere in vivo;unrelated,Raw multiplex: dmsseq AG01046;dmsseq AG01047;dmsseq AG01049;unrelated,,strain:TU/AB|age:2.0;6.0;4.0|dev stage:64c;shield;sphere|sex:pooled male and female|tissue:embryo|treatment:DMS in vivo|molecule:RNA|selection:pA|condition:DMS|replicate group:14;16;15|replicate:2|barcode:CACA;TCTC;AGAG;GTGT|BioSampleModel:Model organism or animal,,,,,,,,,Raw multiplex: DMS Seq   WT 64c in vivo;DMS Seq   WT shield in vivo;DMS Seq   WT sphere in vivo;unrelated,AG01046.3;AG01047.3;AG01049.3;unrelated,AG01046.3;AG01047.3;AG01049.3;unrelated,RNA,,,OTHER,TRANSCRIPTOMIC,unspecified,SINGLE,ILLUMINA,Illumina HiSeq 2500,,SRP114782,,,HA1YVADXX_JBBS71_NOBCX_R1.fastq.gz,fastq,7768733452.0,102220177.0,HA1YVADXX JBBS71 NOBCX R1.fastq.gz,0:76,A:2378019627;C:1963788516;G:1851111018;T:1572409542;N:3404749,76,,,,2378019627,1963788516,1851111018,1572409542,3404749,SRX4168723,SRS3380685,SRA715414,Yale_Giraldez|Genetics,Yale_Giraldez_Group,1,1e-05,,0.0,,1.0,,,,76,,T,,under 1.2% mapping rate,illumina,hiseq_era,unknown,small_rna,unknown,bulk,unknown,unknown,,United States,2018-06-05,Multi-stage,Embryo,Embryo Imprecise,All anatomical structures
43071,SRR7264575,SRX4168722,SRS3380686,SRP114782,PRJNA397065,mRNA structure dynamics identifies RNA remodelers and functional elements during embryogenesis,PRJNA397065,Other,RNA folding plays a crucial role in RNA function. However  our knowledge of the global structure of the transcriptome is limited to steady state conditions  hindering our understanding of how RNA structure dynamics influences gene function. Here  we have characterized mRNA structure dynamics during the maternal to zygotic transition in zebrafish. We observe that on a global level  translation guides structure rather than structure guides translation. We detect a decrease in structure in translated regions  and identify the ribosome as a major remodeler of RNA structure in vivo. In contrast  we find that three prime UTRs form highly folded structures in vivo  which can affect gene expression by modulating miRNA activity. Furthermore  we find that dynamic three prime UTR structures are enriched in RNA decay elements  including regulatory elements in nanog  and cyclin A1  key maternal factors orchestrating the maternal to zygotic transition. These results reveal a central role of RNA structure dynamics in gene regulatory programs during embryogenesis.,,,,Raw multiplex: DMS Seq   WT 64c CHX B1;DMS Seq   WT 64c CHX B2,Raw multiplex: dmsseq AG01273;dmsseq AG01274,,strain:TU/AB|age:2|dev stage:64c|sex:pooled male and female|tissue:embryo|treatment:DMS in vivo cyclohex|molecule:RNA|selection:pA|condition:DMS|replicate group:18|replicate:1;2|barcode:CACA;AGAG|BioSampleModel:Model organism or animal,,,,,,,,,Raw multiplex: DMS Seq   WT 64c CHX B1;DMS Seq   WT 64c CHX B2,AG01273.4;AG01274.4,AG01273.4;AG01274.4,RNA,,,OTHER,TRANSCRIPTOMIC,unspecified,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP114782,,,HWKVWADXX_JBCZ103_023_R1.fastq.gz,fastq,7514617040.0,98876540.0,HWKVWADXX JBCZ103 023 R1.fastq.gz,0:76,A:2401597632;C:1861527917;G:1834777254;T:1415956870;N:757367,76,,,,2401597632,1861527917,1834777254,1415956870,757367,SRX4168722,SRS3380686,SRA715414,Yale_Giraldez|Genetics,Yale_Giraldez_Group,1,0.0,,0.0,,1.0,,,,76,,T,,under 1.2% mapping rate,illumina,hiseq_era,unknown,small_rna,unknown,bulk,unknown,unknown,,United States,2018-06-05,Cleavage,Embryo,Embryo Imprecise,All anatomical structures
43072,SRR7264576,SRX4168721,SRS3380689,SRP114782,PRJNA397065,mRNA structure dynamics identifies RNA remodelers and functional elements during embryogenesis,PRJNA397065,Other,RNA folding plays a crucial role in RNA function. However  our knowledge of the global structure of the transcriptome is limited to steady state conditions  hindering our understanding of how RNA structure dynamics influences gene function. Here  we have characterized mRNA structure dynamics during the maternal to zygotic transition in zebrafish. We observe that on a global level  translation guides structure rather than structure guides translation. We detect a decrease in structure in translated regions  and identify the ribosome as a major remodeler of RNA structure in vivo. In contrast  we find that three prime UTRs form highly folded structures in vivo  which can affect gene expression by modulating miRNA activity. Furthermore  we find that dynamic three prime UTR structures are enriched in RNA decay elements  including regulatory elements in nanog  and cyclin A1  key maternal factors orchestrating the maternal to zygotic transition. These results reveal a central role of RNA structure dynamics in gene regulatory programs during embryogenesis.,,,,Raw multiplex: RPF patA   WT 64c RPF B1;RPF patA   WT 64c RPF B2,Raw multiplex: ribo seq pata AG01409;ribo seq pata AG01410,,strain:TU/AB|age:2|dev stage:64c|sex:pooled male and female|tissue:embryo|treatment:cyclohex|molecule:RNA|condition:RPF 28nt|replicate group:19|replicate:1;2|barcode:CACA;TCTC|BioSampleModel:Model organism or animal,,,,,,,,,Raw multiplex: RPF patA   WT 64c RPF B1;RPF patA   WT 64c RPF B2,AG01409.1;AG01410.1,AG01409.1;AG01410.1,RNA,,,OTHER,TRANSCRIPTOMIC,unspecified,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP114782,,,C7MPPANXX_JBDL115_014_R1.fastq.gz,fastq,4944760108.0,65062633.0,C7MPPANXX JBDL115 014 R1.fastq.gz,0:76,A:1280812305;C:1385761238;G:1441128504;T:836702863;N:355198,76,,,,1280812305,1385761238,1441128504,836702863,355198,SRX4168721,SRS3380689,SRA715414,Yale_Giraldez|Genetics,Yale_Giraldez_Group,1,0.0,,0.0,,1.0,,,,76,,T,,under 1.2% mapping rate,illumina,hiseq_era,unknown,small_rna,unknown,bulk,unknown,unknown,,United States,2018-06-07,Cleavage,Embryo,Embryo Imprecise,All anatomical structures
43073,SRR7264577,SRX4168720,SRS3380688,SRP114782,PRJNA397065,mRNA structure dynamics identifies RNA remodelers and functional elements during embryogenesis,PRJNA397065,Other,RNA folding plays a crucial role in RNA function. However  our knowledge of the global structure of the transcriptome is limited to steady state conditions  hindering our understanding of how RNA structure dynamics influences gene function. Here  we have characterized mRNA structure dynamics during the maternal to zygotic transition in zebrafish. We observe that on a global level  translation guides structure rather than structure guides translation. We detect a decrease in structure in translated regions  and identify the ribosome as a major remodeler of RNA structure in vivo. In contrast  we find that three prime UTRs form highly folded structures in vivo  which can affect gene expression by modulating miRNA activity. Furthermore  we find that dynamic three prime UTR structures are enriched in RNA decay elements  including regulatory elements in nanog  and cyclin A1  key maternal factors orchestrating the maternal to zygotic transition. These results reveal a central role of RNA structure dynamics in gene regulatory programs during embryogenesis.,,,,Raw multiplex: RPF patA   WT 64c PatA RPF B1;RPF patA   WT 64c PatA RPF B2,Raw multiplex: ribo seq pata AG01411;ribo seq pata AG01412,,strain:TU/AB|age:2|dev stage:64c|sex:pooled male and female|tissue:embryo|treatment:cyclohex PatA|molecule:RNA|condition:RPF 28nt|replicate group:20|replicate:1;2|barcode:CACA;TCTC|BioSampleModel:Model organism or animal,,,,,,,,,Raw multiplex: RPF patA   WT 64c PatA RPF B1;RPF patA   WT 64c PatA RPF B2,AG01411.1;AG01412.1,AG01411.1;AG01412.1,RNA,,,OTHER,TRANSCRIPTOMIC,unspecified,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP114782,,,C7MPPANXX_JBDM116_015_R1.fastq.gz,fastq,10995071544.0,144671994.0,C7MPPANXX JBDM116 015 R1.fastq.gz,0:76,A:2850888928;C:3075001734;G:3215965099;T:1852417694;N:798089,76,,,,2850888928,3075001734,3215965099,1852417694,798089,SRX4168720,SRS3380688,SRA715414,Yale_Giraldez|Genetics,Yale_Giraldez_Group,1,0.0,,0.0,,1.0,,,,76,,T,,under 1.2% mapping rate,illumina,hiseq_era,unknown,small_rna,unknown,bulk,unknown,unknown,,United States,2018-06-05,Cleavage,Embryo,Embryo Imprecise,All anatomical structures
43074,SRR7264578,SRX4168719,SRS3380687,SRP114782,PRJNA397065,mRNA structure dynamics identifies RNA remodelers and functional elements during embryogenesis,PRJNA397065,Other,RNA folding plays a crucial role in RNA function. However  our knowledge of the global structure of the transcriptome is limited to steady state conditions  hindering our understanding of how RNA structure dynamics influences gene function. Here  we have characterized mRNA structure dynamics during the maternal to zygotic transition in zebrafish. We observe that on a global level  translation guides structure rather than structure guides translation. We detect a decrease in structure in translated regions  and identify the ribosome as a major remodeler of RNA structure in vivo. In contrast  we find that three prime UTRs form highly folded structures in vivo  which can affect gene expression by modulating miRNA activity. Furthermore  we find that dynamic three prime UTR structures are enriched in RNA decay elements  including regulatory elements in nanog  and cyclin A1  key maternal factors orchestrating the maternal to zygotic transition. These results reveal a central role of RNA structure dynamics in gene regulatory programs during embryogenesis.,,,,Raw multiplex: DMS Seq PatA   WT 64c DMS B1;DMS Seq PatA   WT 64c DMS B2,Raw multiplex: dmsseq pata AG01426;dmsseq pata AG01427,,strain:TU/AB|age:2|dev stage:64c|sex:pooled male and female|tissue:embryo|treatment:DMS|molecule:RNA|selection:pA|condition:DMS|replicate group:21|replicate:1;2|barcode:CACA;AGAG|BioSampleModel:Model organism or animal,,,,,,,,,Raw multiplex: DMS Seq PatA   WT 64c DMS B1;DMS Seq PatA   WT 64c DMS B2,AG01426.1;AG01427.1,AG01426.1;AG01427.1,RNA,,,OTHER,TRANSCRIPTOMIC,unspecified,SINGLE,ILLUMINA,Illumina HiSeq 2000,,SRP114782,,,C7MPPANXX_JBDF109_027_R1.fastq.gz,fastq,446991948.0,5881473.0,C7MPPANXX JBDF109 027 R1.fastq.gz,0:76,A:146192524;C:109441919;G:103882304;T:87445288;N:29913,76,,,,146192524,109441919,103882304,87445288,29913,SRX4168719,SRS3380687,SRA715414,Yale_Giraldez|Genetics,Yale_Giraldez_Group,1,2e-05,,1e-05,,1.0,,,,76,,T,,under 1.2% mapping rate,illumina,hiseq_era,unknown,small_rna,unknown,bulk,unknown,unknown,,United States,2018-06-07,Cleavage,Embryo,Embryo Imprecise,All anatomical structures