{"database": "metadata", "table": "run_metadata", "is_view": false, "human_description_en": "where devstage_curation = \"Zygote\" and experiment.library_selection = \"other\"", "rows": [[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.", null, null, null, "Fish embryo mRNA 1024cell PAL seq v4", "GSM7716868", null, "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 \u201c m 15 quality base=64 q 20 20 match read wildcards e 0.05 a NNNNATCTCGTATGCCGTCTTCTGCTTG O 7\u201d. 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 \u201c runThreadN 16 runMode alignReads outFilterMultimapNmax 1 outReadsUnmapped Fastx outFilterType BySJout outSAMattributes All outSAMtype BAM Unsorted SortedByCoordinate\u201d. 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\u20137 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", null, "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 \u00b5g/ml cycloheximide cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer] and 200 units/ml SUPERase\u2022In in a volume of 10 \u00b5l per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4\u00b0C 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\u00b0C 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 3\u02b9 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 \u00b5l. The RNA and oligos were annealed by incubation at 70\u00b0C for 5 min and then slowly cooling to 23\u00b0C at 0.1\u00b0C/sec. The annealed mixture was combined with 40 \u00b5l MyOne Streptavidin C1 beads Thermo Fisher 65002 and incubated for 20 min at 23\u00b0C 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 \u00b5l 1xB&W buffer 5 mM Tris HCl pH 7.5 0.5 mM EDTA 1 M NaCl and once with 300 \u00b5l 2x SSC. The RNA was eluted from the beads first with 100 \u00b5l 10 mM HEPES pH 7.5 at 65\u00b0C for 3 min and then second with 100 \u00b5l water at 65\u00b0C for 3 min. The eluates were combined precipitated with ethanol and resuspended in 6.5 \u00b5l water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3\u02b9 adapter in a 10 \u00b5l reaction containing 5 \u00b5M 3\u02b9 adapter KXS330 50 mM HEPES pH 7.5 10 mM MgCl2 10 mM dithiothreitol 1 unit/\u00b5l T4 RNA ligase 1 New England Biolabs M0204S. The ligation reaction was incubated at 23\u00b0C for 150 min. post ligation RNA was extracted with phenol/chloroform precipitated with ethanol and resuspended in 11.4 \u00b5l water. The ligated RNA was mixed with 0.6 \u00b5l 100 \u00b5M reverse transcription primer KXS037 in a total volume of 12 \u00b5l incubated at 65\u00b0C for 5 min and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 \u00b5l reaction containing 1x First Strand Buffer 500 \u00b5M dNTPs 5 mM dithiothreitol 1 unit/\u00b5l SUPERase\u2022In and 200 units SuperScript III Thermo Fisher 18080044 at 50\u00b0C for 1 hr. post reverse transcription RNA was hydrolyzed with 3.3 \u00b5l 1 M NaOH at 90\u00b0C for 10 min followed by neutralization with 36.7 \u00b5l 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 \u00b5l PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer\u2019s suggested protocol for 10\u201315 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 3\u02b9 adapter KXS013 was used for 3\u02b9 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 \u00b5l. The cDNAs and the oligo were annealed by incubation at 65\u00b0C for 5 min and then slowly cooling to 23\u00b0C at 0.1\u00b0C/sec. The annealed mixture was combined with 100 \u00b5l MyOne Streptavidin C1 beads and incubated for 20 min at 23\u00b0C 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 \u00b5l 1xB&W buffer. The supernatant and the wash were combined precipitated with ethanol and resuspended in 30 \u00b5l water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.", null, "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 \u00b5g/ml cycloheximide cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer] and 200 units/ml SUPERase\u2022In in a volume of 10 \u00b5l per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4\u00b0C 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\u00b0C 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 3\u02b9 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 \u00b5l. The RNA and oligos were annealed by incubation at 70\u00b0C for 5 min and then slowly cooling to 23\u00b0C at 0.1\u00b0C/sec. The annealed mixture was combined with 40 \u00b5l MyOne Streptavidin C1 beads Thermo Fisher 65002 and incubated for 20 min at 23\u00b0C 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 \u00b5l 1xB&W buffer 5 mM Tris HCl pH 7.5 0.5 mM EDTA 1 M NaCl and once with 300 \u00b5l 2x SSC. The RNA was eluted from the beads first with 100 \u00b5l 10 mM HEPES pH 7.5 at 65\u00b0C for 3 min and then second with 100 \u00b5l water at 65\u00b0C for 3 min. The eluates were combined precipitated with ethanol and resuspended in 6.5 \u00b5l water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3\u02b9 adapter in a 10 \u00b5l reaction containing 5 \u00b5M 3\u02b9 adapter KXS330 50 mM HEPES pH 7.5 10 mM MgCl2 10 mM dithiothreitol 1 unit/\u00b5l T4 RNA ligase 1 New England Biolabs M0204S. The ligation reaction was incubated at 23\u00b0C for 150 min. post ligation RNA was extracted with phenol/chloroform precipitated with ethanol and resuspended in 11.4 \u00b5l water. The ligated RNA was mixed with 0.6 \u00b5l 100 \u00b5M reverse transcription primer KXS037 in a total volume of 12 \u00b5l incubated at 65\u00b0C for 5 min and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 \u00b5l reaction containing 1x First Strand Buffer 500 \u00b5M dNTPs 5 mM dithiothreitol 1 unit/\u00b5l SUPERase\u2022In and 200 units SuperScript III Thermo Fisher 18080044 at 50\u00b0C for 1 hr. post reverse transcription RNA was hydrolyzed with 3.3 \u00b5l 1 M NaOH at 90\u00b0C for 10 min followed by neutralization with 36.7 \u00b5l 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 \u00b5l PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer's suggested protocol for 10\u201315 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 3\u02b9 adapter KXS013 was used for 3\u02b9 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 \u00b5l. The cDNAs and the oligo were annealed by incubation at 65\u00b0C for 5 min and then slowly cooling to 23\u00b0C at 0.1\u00b0C/sec. The annealed mixture was combined with 100 \u00b5l MyOne Streptavidin C1 beads and incubated for 20 min at 23\u00b0C 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 \u00b5l 1xB&W buffer. The supernatant and the wash were combined precipitated with ethanol and resuspended in 30 \u00b5l water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.", null, "OTHER", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP455680", null, null, "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, null, null, 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", null, "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.", null, null, null, "Fish embryo mRNA 1024cell PAL seq v4", "GSM7716868", null, "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 \u201c m 15 quality base=64 q 20 20 match read wildcards e 0.05 a NNNNATCTCGTATGCCGTCTTCTGCTTG O 7\u201d. 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 \u201c runThreadN 16 runMode alignReads outFilterMultimapNmax 1 outReadsUnmapped Fastx outFilterType BySJout outSAMattributes All outSAMtype BAM Unsorted SortedByCoordinate\u201d. 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\u20137 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", null, "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 \u00b5g/ml cycloheximide cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer] and 200 units/ml SUPERase\u2022In in a volume of 10 \u00b5l per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4\u00b0C 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\u00b0C 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 3\u02b9 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 \u00b5l. The RNA and oligos were annealed by incubation at 70\u00b0C for 5 min and then slowly cooling to 23\u00b0C at 0.1\u00b0C/sec. The annealed mixture was combined with 40 \u00b5l MyOne Streptavidin C1 beads Thermo Fisher 65002 and incubated for 20 min at 23\u00b0C 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 \u00b5l 1xB&W buffer 5 mM Tris HCl pH 7.5 0.5 mM EDTA 1 M NaCl and once with 300 \u00b5l 2x SSC. The RNA was eluted from the beads first with 100 \u00b5l 10 mM HEPES pH 7.5 at 65\u00b0C for 3 min and then second with 100 \u00b5l water at 65\u00b0C for 3 min. The eluates were combined precipitated with ethanol and resuspended in 6.5 \u00b5l water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3\u02b9 adapter in a 10 \u00b5l reaction containing 5 \u00b5M 3\u02b9 adapter KXS330 50 mM HEPES pH 7.5 10 mM MgCl2 10 mM dithiothreitol 1 unit/\u00b5l T4 RNA ligase 1 New England Biolabs M0204S. The ligation reaction was incubated at 23\u00b0C for 150 min. post ligation RNA was extracted with phenol/chloroform precipitated with ethanol and resuspended in 11.4 \u00b5l water. The ligated RNA was mixed with 0.6 \u00b5l 100 \u00b5M reverse transcription primer KXS037 in a total volume of 12 \u00b5l incubated at 65\u00b0C for 5 min and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 \u00b5l reaction containing 1x First Strand Buffer 500 \u00b5M dNTPs 5 mM dithiothreitol 1 unit/\u00b5l SUPERase\u2022In and 200 units SuperScript III Thermo Fisher 18080044 at 50\u00b0C for 1 hr. post reverse transcription RNA was hydrolyzed with 3.3 \u00b5l 1 M NaOH at 90\u00b0C for 10 min followed by neutralization with 36.7 \u00b5l 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 \u00b5l PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer\u2019s suggested protocol for 10\u201315 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 3\u02b9 adapter KXS013 was used for 3\u02b9 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 \u00b5l. The cDNAs and the oligo were annealed by incubation at 65\u00b0C for 5 min and then slowly cooling to 23\u00b0C at 0.1\u00b0C/sec. The annealed mixture was combined with 100 \u00b5l MyOne Streptavidin C1 beads and incubated for 20 min at 23\u00b0C 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 \u00b5l 1xB&W buffer. The supernatant and the wash were combined precipitated with ethanol and resuspended in 30 \u00b5l water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.", null, "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 \u00b5g/ml cycloheximide cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer] and 200 units/ml SUPERase\u2022In in a volume of 10 \u00b5l per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4\u00b0C 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\u00b0C 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 3\u02b9 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 \u00b5l. The RNA and oligos were annealed by incubation at 70\u00b0C for 5 min and then slowly cooling to 23\u00b0C at 0.1\u00b0C/sec. The annealed mixture was combined with 40 \u00b5l MyOne Streptavidin C1 beads Thermo Fisher 65002 and incubated for 20 min at 23\u00b0C 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 \u00b5l 1xB&W buffer 5 mM Tris HCl pH 7.5 0.5 mM EDTA 1 M NaCl and once with 300 \u00b5l 2x SSC. The RNA was eluted from the beads first with 100 \u00b5l 10 mM HEPES pH 7.5 at 65\u00b0C for 3 min and then second with 100 \u00b5l water at 65\u00b0C for 3 min. The eluates were combined precipitated with ethanol and resuspended in 6.5 \u00b5l water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3\u02b9 adapter in a 10 \u00b5l reaction containing 5 \u00b5M 3\u02b9 adapter KXS330 50 mM HEPES pH 7.5 10 mM MgCl2 10 mM dithiothreitol 1 unit/\u00b5l T4 RNA ligase 1 New England Biolabs M0204S. The ligation reaction was incubated at 23\u00b0C for 150 min. post ligation RNA was extracted with phenol/chloroform precipitated with ethanol and resuspended in 11.4 \u00b5l water. The ligated RNA was mixed with 0.6 \u00b5l 100 \u00b5M reverse transcription primer KXS037 in a total volume of 12 \u00b5l incubated at 65\u00b0C for 5 min and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 \u00b5l reaction containing 1x First Strand Buffer 500 \u00b5M dNTPs 5 mM dithiothreitol 1 unit/\u00b5l SUPERase\u2022In and 200 units SuperScript III Thermo Fisher 18080044 at 50\u00b0C for 1 hr. post reverse transcription RNA was hydrolyzed with 3.3 \u00b5l 1 M NaOH at 90\u00b0C for 10 min followed by neutralization with 36.7 \u00b5l 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 \u00b5l PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer's suggested protocol for 10\u201315 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 3\u02b9 adapter KXS013 was used for 3\u02b9 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 \u00b5l. The cDNAs and the oligo were annealed by incubation at 65\u00b0C for 5 min and then slowly cooling to 23\u00b0C at 0.1\u00b0C/sec. The annealed mixture was combined with 100 \u00b5l MyOne Streptavidin C1 beads and incubated for 20 min at 23\u00b0C 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 \u00b5l 1xB&W buffer. The supernatant and the wash were combined precipitated with ethanol and resuspended in 30 \u00b5l water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.", null, "OTHER", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP455680", null, null, "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, null, null, 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, null, 52, 255, "T", "T", "mates < 9% mapping rate", "illumina", "hiseq_era", "unknown", "poly_a", "unknown", "bulk", "bulk", "bulk", null, "United States", "2023-08-17", "Zygote", "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.", null, null, null, "Fish embryo mRNA 1cell PAL seq v4", "GSM7716865", null, "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 \u201c m 15 quality base=64 q 20 20 match read wildcards e 0.05 a NNNNATCTCGTATGCCGTCTTCTGCTTG O 7\u201d. 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 \u201c runThreadN 16 runMode alignReads outFilterMultimapNmax 1 outReadsUnmapped Fastx outFilterType BySJout outSAMattributes All outSAMtype BAM Unsorted SortedByCoordinate\u201d. 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\u20137 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", null, "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 \u00b5g/ml cycloheximide cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer] and 200 units/ml SUPERase\u2022In in a volume of 10 \u00b5l per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4\u00b0C 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\u00b0C 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 3\u02b9 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 \u00b5l. The RNA and oligos were annealed by incubation at 70\u00b0C for 5 min and then slowly cooling to 23\u00b0C at 0.1\u00b0C/sec. The annealed mixture was combined with 40 \u00b5l MyOne Streptavidin C1 beads Thermo Fisher 65002 and incubated for 20 min at 23\u00b0C 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 \u00b5l 1xB&W buffer 5 mM Tris HCl pH 7.5 0.5 mM EDTA 1 M NaCl and once with 300 \u00b5l 2x SSC. The RNA was eluted from the beads first with 100 \u00b5l 10 mM HEPES pH 7.5 at 65\u00b0C for 3 min and then second with 100 \u00b5l water at 65\u00b0C for 3 min. The eluates were combined precipitated with ethanol and resuspended in 6.5 \u00b5l water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3\u02b9 adapter in a 10 \u00b5l reaction containing 5 \u00b5M 3\u02b9 adapter KXS330 50 mM HEPES pH 7.5 10 mM MgCl2 10 mM dithiothreitol 1 unit/\u00b5l T4 RNA ligase 1 New England Biolabs M0204S. The ligation reaction was incubated at 23\u00b0C for 150 min. post ligation RNA was extracted with phenol/chloroform precipitated with ethanol and resuspended in 11.4 \u00b5l water. The ligated RNA was mixed with 0.6 \u00b5l 100 \u00b5M reverse transcription primer KXS037 in a total volume of 12 \u00b5l incubated at 65\u00b0C for 5 min and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 \u00b5l reaction containing 1x First Strand Buffer 500 \u00b5M dNTPs 5 mM dithiothreitol 1 unit/\u00b5l SUPERase\u2022In and 200 units SuperScript III Thermo Fisher 18080044 at 50\u00b0C for 1 hr. post reverse transcription RNA was hydrolyzed with 3.3 \u00b5l 1 M NaOH at 90\u00b0C for 10 min followed by neutralization with 36.7 \u00b5l 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 \u00b5l PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer\u2019s suggested protocol for 10\u201315 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 3\u02b9 adapter KXS013 was used for 3\u02b9 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 \u00b5l. The cDNAs and the oligo were annealed by incubation at 65\u00b0C for 5 min and then slowly cooling to 23\u00b0C at 0.1\u00b0C/sec. The annealed mixture was combined with 100 \u00b5l MyOne Streptavidin C1 beads and incubated for 20 min at 23\u00b0C 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 \u00b5l 1xB&W buffer. The supernatant and the wash were combined precipitated with ethanol and resuspended in 30 \u00b5l water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.", null, "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 \u00b5g/ml cycloheximide cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer] and 200 units/ml SUPERase\u2022In in a volume of 10 \u00b5l per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4\u00b0C 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\u00b0C 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 3\u02b9 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 \u00b5l. The RNA and oligos were annealed by incubation at 70\u00b0C for 5 min and then slowly cooling to 23\u00b0C at 0.1\u00b0C/sec. The annealed mixture was combined with 40 \u00b5l MyOne Streptavidin C1 beads Thermo Fisher 65002 and incubated for 20 min at 23\u00b0C 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 \u00b5l 1xB&W buffer 5 mM Tris HCl pH 7.5 0.5 mM EDTA 1 M NaCl and once with 300 \u00b5l 2x SSC. The RNA was eluted from the beads first with 100 \u00b5l 10 mM HEPES pH 7.5 at 65\u00b0C for 3 min and then second with 100 \u00b5l water at 65\u00b0C for 3 min. The eluates were combined precipitated with ethanol and resuspended in 6.5 \u00b5l water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3\u02b9 adapter in a 10 \u00b5l reaction containing 5 \u00b5M 3\u02b9 adapter KXS330 50 mM HEPES pH 7.5 10 mM MgCl2 10 mM dithiothreitol 1 unit/\u00b5l T4 RNA ligase 1 New England Biolabs M0204S. The ligation reaction was incubated at 23\u00b0C for 150 min. post ligation RNA was extracted with phenol/chloroform precipitated with ethanol and resuspended in 11.4 \u00b5l water. The ligated RNA was mixed with 0.6 \u00b5l 100 \u00b5M reverse transcription primer KXS037 in a total volume of 12 \u00b5l incubated at 65\u00b0C for 5 min and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 \u00b5l reaction containing 1x First Strand Buffer 500 \u00b5M dNTPs 5 mM dithiothreitol 1 unit/\u00b5l SUPERase\u2022In and 200 units SuperScript III Thermo Fisher 18080044 at 50\u00b0C for 1 hr. post reverse transcription RNA was hydrolyzed with 3.3 \u00b5l 1 M NaOH at 90\u00b0C for 10 min followed by neutralization with 36.7 \u00b5l 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 \u00b5l PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer's suggested protocol for 10\u201315 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 3\u02b9 adapter KXS013 was used for 3\u02b9 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 \u00b5l. The cDNAs and the oligo were annealed by incubation at 65\u00b0C for 5 min and then slowly cooling to 23\u00b0C at 0.1\u00b0C/sec. The annealed mixture was combined with 100 \u00b5l MyOne Streptavidin C1 beads and incubated for 20 min at 23\u00b0C 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 \u00b5l 1xB&W buffer. The supernatant and the wash were combined precipitated with ethanol and resuspended in 30 \u00b5l water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.", null, "OTHER", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP455680", null, null, "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, null, null, 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", null, "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.", null, null, null, "Fish embryo mRNA 1cell PAL seq v4", "GSM7716865", null, "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 \u201c m 15 quality base=64 q 20 20 match read wildcards e 0.05 a NNNNATCTCGTATGCCGTCTTCTGCTTG O 7\u201d. 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 \u201c runThreadN 16 runMode alignReads outFilterMultimapNmax 1 outReadsUnmapped Fastx outFilterType BySJout outSAMattributes All outSAMtype BAM Unsorted SortedByCoordinate\u201d. 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\u20137 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", null, "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 \u00b5g/ml cycloheximide cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer] and 200 units/ml SUPERase\u2022In in a volume of 10 \u00b5l per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4\u00b0C 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\u00b0C 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 3\u02b9 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 \u00b5l. The RNA and oligos were annealed by incubation at 70\u00b0C for 5 min and then slowly cooling to 23\u00b0C at 0.1\u00b0C/sec. The annealed mixture was combined with 40 \u00b5l MyOne Streptavidin C1 beads Thermo Fisher 65002 and incubated for 20 min at 23\u00b0C 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 \u00b5l 1xB&W buffer 5 mM Tris HCl pH 7.5 0.5 mM EDTA 1 M NaCl and once with 300 \u00b5l 2x SSC. The RNA was eluted from the beads first with 100 \u00b5l 10 mM HEPES pH 7.5 at 65\u00b0C for 3 min and then second with 100 \u00b5l water at 65\u00b0C for 3 min. The eluates were combined precipitated with ethanol and resuspended in 6.5 \u00b5l water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3\u02b9 adapter in a 10 \u00b5l reaction containing 5 \u00b5M 3\u02b9 adapter KXS330 50 mM HEPES pH 7.5 10 mM MgCl2 10 mM dithiothreitol 1 unit/\u00b5l T4 RNA ligase 1 New England Biolabs M0204S. The ligation reaction was incubated at 23\u00b0C for 150 min. post ligation RNA was extracted with phenol/chloroform precipitated with ethanol and resuspended in 11.4 \u00b5l water. The ligated RNA was mixed with 0.6 \u00b5l 100 \u00b5M reverse transcription primer KXS037 in a total volume of 12 \u00b5l incubated at 65\u00b0C for 5 min and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 \u00b5l reaction containing 1x First Strand Buffer 500 \u00b5M dNTPs 5 mM dithiothreitol 1 unit/\u00b5l SUPERase\u2022In and 200 units SuperScript III Thermo Fisher 18080044 at 50\u00b0C for 1 hr. post reverse transcription RNA was hydrolyzed with 3.3 \u00b5l 1 M NaOH at 90\u00b0C for 10 min followed by neutralization with 36.7 \u00b5l 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 \u00b5l PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer\u2019s suggested protocol for 10\u201315 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 3\u02b9 adapter KXS013 was used for 3\u02b9 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 \u00b5l. The cDNAs and the oligo were annealed by incubation at 65\u00b0C for 5 min and then slowly cooling to 23\u00b0C at 0.1\u00b0C/sec. The annealed mixture was combined with 100 \u00b5l MyOne Streptavidin C1 beads and incubated for 20 min at 23\u00b0C 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 \u00b5l 1xB&W buffer. The supernatant and the wash were combined precipitated with ethanol and resuspended in 30 \u00b5l water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.", null, "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 \u00b5g/ml cycloheximide cOmplete protease inhibitor cocktail [1 tablet per 10 ml buffer] and 200 units/ml SUPERase\u2022In in a volume of 10 \u00b5l per oocyte/embryo by vigorous shaking and pipetting. Lysates were cleared by centrifugation at 5000 g at 4\u00b0C 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\u00b0C 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 3\u02b9 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 \u00b5l. The RNA and oligos were annealed by incubation at 70\u00b0C for 5 min and then slowly cooling to 23\u00b0C at 0.1\u00b0C/sec. The annealed mixture was combined with 40 \u00b5l MyOne Streptavidin C1 beads Thermo Fisher 65002 and incubated for 20 min at 23\u00b0C 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 \u00b5l 1xB&W buffer 5 mM Tris HCl pH 7.5 0.5 mM EDTA 1 M NaCl and once with 300 \u00b5l 2x SSC. The RNA was eluted from the beads first with 100 \u00b5l 10 mM HEPES pH 7.5 at 65\u00b0C for 3 min and then second with 100 \u00b5l water at 65\u00b0C for 3 min. The eluates were combined precipitated with ethanol and resuspended in 6.5 \u00b5l water. The anti sense oligo enriched RNA or the RNA isolated from oocyte or embryo lysates was ligated to a pre adenylated 3\u02b9 adapter in a 10 \u00b5l reaction containing 5 \u00b5M 3\u02b9 adapter KXS330 50 mM HEPES pH 7.5 10 mM MgCl2 10 mM dithiothreitol 1 unit/\u00b5l T4 RNA ligase 1 New England Biolabs M0204S. The ligation reaction was incubated at 23\u00b0C for 150 min. post ligation RNA was extracted with phenol/chloroform precipitated with ethanol and resuspended in 11.4 \u00b5l water. The ligated RNA was mixed with 0.6 \u00b5l 100 \u00b5M reverse transcription primer KXS037 in a total volume of 12 \u00b5l incubated at 65\u00b0C for 5 min and cooled on ice for 1 min. The annealed RNA was reverse transcribed in a 20 \u00b5l reaction containing 1x First Strand Buffer 500 \u00b5M dNTPs 5 mM dithiothreitol 1 unit/\u00b5l SUPERase\u2022In and 200 units SuperScript III Thermo Fisher 18080044 at 50\u00b0C for 1 hr. post reverse transcription RNA was hydrolyzed with 3.3 \u00b5l 1 M NaOH at 90\u00b0C for 10 min followed by neutralization with 36.7 \u00b5l 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 \u00b5l PCR reaction with KXS037 and a barcoded primer Supplemental using the KAPA HiFi HotStart Kits following the manufacturer's suggested protocol for 10\u201315 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 3\u02b9 adapter KXS013 was used for 3\u02b9 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 \u00b5l. The cDNAs and the oligo were annealed by incubation at 65\u00b0C for 5 min and then slowly cooling to 23\u00b0C at 0.1\u00b0C/sec. The annealed mixture was combined with 100 \u00b5l MyOne Streptavidin C1 beads and incubated for 20 min at 23\u00b0C 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 \u00b5l 1xB&W buffer. The supernatant and the wash were combined precipitated with ethanol and resuspended in 30 \u00b5l water. The oligo depleted libraries were sequenced again as a technical replicate. Sequencing data of replicates were merged for each sample post monitoring consistency.", null, "OTHER", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP455680", null, null, "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, null, null, 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, null, 52, 255, "T", "T", "mates < 9% mapping rate", "illumina", "hiseq_era", "unknown", "poly_a", "unknown", "bulk", "bulk", "bulk", null, "United States", "2023-08-17", "Zygote", "Embryo", "Embryo Imprecise", "All anatomical structures"], [40701, "SRR3231363", "SRX1637111", "SRS1342926", "SRP071849", "PRJNA315400", "Tdrd6a recruits Ziwi bound piRNAs and coordinates deposition of germ plasm mRNAs into primordial germ cells [RIP Seq]", "GSE79161", "Transcriptome Analysis", "Germ plasm the Balbiani body and nuage are evolutionary conserved structures essential for germ cell specification and maintenance. We describe Tdrd6a as a component of these structures with two distinct molecular functions. First Tdrd6a facilitates the accumulation of the typical antisense bias of piRNAs without xxx effects on piRNA biogenesis signatures. Second we show that Tdrd6a is required for Balbiani body and germ plasm integrity and associates with RNA binding proteins and germ plasm mRNAs. On the cell biological level maternally contributed Tdrd6a strongly impacts germ cell formation but is dispensable for fertility. Using single cell RNA sequencing we demonstrate that Tdrd6a promotes early germ cell development and regulates the stoichiometry of germ plasm mRNAs. We propose that Tdrd6a functions as a scaffold to recruit correct ratios of germ plasm transcripts and to accumulate antisense piRNA complexes in order to ensure both specification and maintenance of germ cells. Overall design: RNA was extracted from ovary tissue or from RIP experiments by Trizol extraction.", "parent bioproject:PRJNA315403", "pubmed:30086300", null, "Input 3", "GSM2087141", null, "tissue:Zebrafish Zygotes|fraction:Input", "Input 3", "Reads were mapped to Zv9 using tophat trapnell et al 2009to the ENSEMBLE gene build and further processing of the read counts was performed using Deseq Anders et al 2009 Genome build: zv9 Supplementary files format and content: The mRNA counts.csv contains the DESeq normalized read counts per Ensemble gene as indicated. Headers indicate the sample name.", "Zebrafish Zygotes", null, "mRNA was extracted from zygotes or by RIP experiments followed by Trizol extraction The RNA was extracted using chloroform and precipitated with iso propanol. Ovation RNA seq System V2 NuGEN library preperation kit was used and the library was sequenced on an Illumina HISEQ using 50bp single end sequencing.", "Zebrafish were maintained under standard conditions.", "fraction:Input", "GSM2087141", "GSM2087141: Input 3; Danio rerio; RIP Seq", "GSM2087141", null, "1", "mRNA was extracted from zygotes or by RIP experiments followed by Trizol extraction The RNA was extracted using chloroform and precipitated with iso propanol. Ovation RNA seq System V2 NuGEN library preperation kit was used and the library was sequenced on an Illumina HISEQ using 50bp single end sequencing.", "GEO Accession:GSM2087141", "RIP-Seq", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2000", null, "SRP071849", null, null, "WT_Input_3.fastq.gz", "fastq", 1902296226.0, 37299926.0, "GSM2087141 r1", "0:51", "A:466290166;C:465344039;G:486942278;T:483240280;N:479463", 51, null, null, null, 466290166, 465344039, 486942278, 483240280, 479463, "SRX1637111", "SRS1342926", "SRA385813", "GEO", "Rene Ketting, RNA silencing, IMB", 1, 0.85793, null, 0.14814, null, 0.79941, null, 0.55214, null, 51, null, "B", null, "usable mapping rate", "illumina", "hiseq_era", "unknown", "other", "unknown", "sc_generic", "single_cell_generic", "generic-scrnaseq-only", null, "Germany", "2016-03-13", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [40702, "SRR3231362", "SRX1637110", "SRS1342927", "SRP071849", "PRJNA315400", "Tdrd6a recruits Ziwi bound piRNAs and coordinates deposition of germ plasm mRNAs into primordial germ cells [RIP Seq]", "GSE79161", "Transcriptome Analysis", "Germ plasm the Balbiani body and nuage are evolutionary conserved structures essential for germ cell specification and maintenance. We describe Tdrd6a as a component of these structures with two distinct molecular functions. First Tdrd6a facilitates the accumulation of the typical antisense bias of piRNAs without xxx effects on piRNA biogenesis signatures. Second we show that Tdrd6a is required for Balbiani body and germ plasm integrity and associates with RNA binding proteins and germ plasm mRNAs. On the cell biological level maternally contributed Tdrd6a strongly impacts germ cell formation but is dispensable for fertility. Using single cell RNA sequencing we demonstrate that Tdrd6a promotes early germ cell development and regulates the stoichiometry of germ plasm mRNAs. We propose that Tdrd6a functions as a scaffold to recruit correct ratios of germ plasm transcripts and to accumulate antisense piRNA complexes in order to ensure both specification and maintenance of germ cells. Overall design: RNA was extracted from ovary tissue or from RIP experiments by Trizol extraction.", "parent bioproject:PRJNA315403", "pubmed:30086300", null, "Input 2", "GSM2087140", null, "tissue:Zebrafish Zygotes|fraction:Input", "Input 2", "Reads were mapped to Zv9 using tophat trapnell et al 2009to the ENSEMBLE gene build and further processing of the read counts was performed using Deseq Anders et al 2009 Genome build: zv9 Supplementary files format and content: The mRNA counts.csv contains the DESeq normalized read counts per Ensemble gene as indicated. Headers indicate the sample name.", "Zebrafish Zygotes", null, "mRNA was extracted from zygotes or by RIP experiments followed by Trizol extraction The RNA was extracted using chloroform and precipitated with iso propanol. Ovation RNA seq System V2 NuGEN library preperation kit was used and the library was sequenced on an Illumina HISEQ using 50bp single end sequencing.", "Zebrafish were maintained under standard conditions.", "fraction:Input", "GSM2087140", "GSM2087140: Input 2; Danio rerio; RIP Seq", "GSM2087140", null, "1", "mRNA was extracted from zygotes or by RIP experiments followed by Trizol extraction The RNA was extracted using chloroform and precipitated with iso propanol. Ovation RNA seq System V2 NuGEN library preperation kit was used and the library was sequenced on an Illumina HISEQ using 50bp single end sequencing.", "GEO Accession:GSM2087140", "RIP-Seq", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2000", null, "SRP071849", null, null, "WT_Input_2.fastq.gz", "fastq", 1540737999.0, 30210549.0, "GSM2087140 r1", "0:51", "A:380481772;C:379741192;G:395453141;T:384675313;N:386581", 51, null, null, null, 380481772, 379741192, 395453141, 384675313, 386581, "SRX1637110", "SRS1342927", "SRA385813", "GEO", "Rene Ketting, RNA silencing, IMB", 1, 0.8787, null, 0.14935, null, 0.79202, null, 0.54381, null, 51, null, "B", null, "usable mapping rate", "illumina", "hiseq_era", "unknown", "other", "unknown", "sc_generic", "single_cell_generic", "generic-scrnaseq-only", null, "Germany", "2016-03-13", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [40703, "SRR3231361", "SRX1637109", "SRS1342928", "SRP071849", "PRJNA315400", "Tdrd6a recruits Ziwi bound piRNAs and coordinates deposition of germ plasm mRNAs into primordial germ cells [RIP Seq]", "GSE79161", "Transcriptome Analysis", "Germ plasm the Balbiani body and nuage are evolutionary conserved structures essential for germ cell specification and maintenance. We describe Tdrd6a as a component of these structures with two distinct molecular functions. First Tdrd6a facilitates the accumulation of the typical antisense bias of piRNAs without xxx effects on piRNA biogenesis signatures. Second we show that Tdrd6a is required for Balbiani body and germ plasm integrity and associates with RNA binding proteins and germ plasm mRNAs. On the cell biological level maternally contributed Tdrd6a strongly impacts germ cell formation but is dispensable for fertility. Using single cell RNA sequencing we demonstrate that Tdrd6a promotes early germ cell development and regulates the stoichiometry of germ plasm mRNAs. We propose that Tdrd6a functions as a scaffold to recruit correct ratios of germ plasm transcripts and to accumulate antisense piRNA complexes in order to ensure both specification and maintenance of germ cells. Overall design: RNA was extracted from ovary tissue or from RIP experiments by Trizol extraction.", "parent bioproject:PRJNA315403", "pubmed:30086300", null, "Input 1", "GSM2087139", null, "tissue:Zebrafish Zygotes|fraction:Input", "Input 1", "Reads were mapped to Zv9 using tophat trapnell et al 2009to the ENSEMBLE gene build and further processing of the read counts was performed using Deseq Anders et al 2009 Genome build: zv9 Supplementary files format and content: The mRNA counts.csv contains the DESeq normalized read counts per Ensemble gene as indicated. Headers indicate the sample name.", "Zebrafish Zygotes", null, "mRNA was extracted from zygotes or by RIP experiments followed by Trizol extraction The RNA was extracted using chloroform and precipitated with iso propanol. Ovation RNA seq System V2 NuGEN library preperation kit was used and the library was sequenced on an Illumina HISEQ using 50bp single end sequencing.", "Zebrafish were maintained under standard conditions.", "fraction:Input", "GSM2087139", "GSM2087139: Input 1; Danio rerio; RIP Seq", "GSM2087139", null, "1", "mRNA was extracted from zygotes or by RIP experiments followed by Trizol extraction The RNA was extracted using chloroform and precipitated with iso propanol. Ovation RNA seq System V2 NuGEN library preperation kit was used and the library was sequenced on an Illumina HISEQ using 50bp single end sequencing.", "GEO Accession:GSM2087139", "RIP-Seq", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2000", null, "SRP071849", null, null, "WT_Input_1.fastq.gz", "fastq", 1109818905.0, 21761155.0, "GSM2087139 r1", null, null, null, null, null, null, null, null, null, null, null, "SRX1637109", "SRS1342928", "SRA385813", "GEO", "Rene Ketting, RNA silencing, IMB", 1, 0.87467, null, 0.18794, null, 0.79411, null, 0.63378, null, 51, null, "B", null, "usable mapping rate", "illumina", "hiseq_era", "unknown", "other", "unknown", "sc_generic", "single_cell_generic", "generic-scrnaseq-only", null, "Germany", "2016-03-13", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [40704, "SRR3231360", "SRX1637108", "SRS1342929", "SRP071849", "PRJNA315400", "Tdrd6a recruits Ziwi bound piRNAs and coordinates deposition of germ plasm mRNAs into primordial germ cells [RIP Seq]", "GSE79161", "Transcriptome Analysis", "Germ plasm the Balbiani body and nuage are evolutionary conserved structures essential for germ cell specification and maintenance. We describe Tdrd6a as a component of these structures with two distinct molecular functions. First Tdrd6a facilitates the accumulation of the typical antisense bias of piRNAs without xxx effects on piRNA biogenesis signatures. Second we show that Tdrd6a is required for Balbiani body and germ plasm integrity and associates with RNA binding proteins and germ plasm mRNAs. On the cell biological level maternally contributed Tdrd6a strongly impacts germ cell formation but is dispensable for fertility. Using single cell RNA sequencing we demonstrate that Tdrd6a promotes early germ cell development and regulates the stoichiometry of germ plasm mRNAs. We propose that Tdrd6a functions as a scaffold to recruit correct ratios of germ plasm transcripts and to accumulate antisense piRNA complexes in order to ensure both specification and maintenance of germ cells. Overall design: RNA was extracted from ovary tissue or from RIP experiments by Trizol extraction.", "parent bioproject:PRJNA315403", "pubmed:30086300", null, "Tdrd6a IP 3", "GSM2087138", null, "tissue:Zebrafish Zygotes|fraction:Tdrd6a IP", "Tdrd6a IP 3", "Reads were mapped to Zv9 using tophat trapnell et al 2009to the ENSEMBLE gene build and further processing of the read counts was performed using Deseq Anders et al 2009 Genome build: zv9 Supplementary files format and content: The mRNA counts.csv contains the DESeq normalized read counts per Ensemble gene as indicated. Headers indicate the sample name.", "Zebrafish Zygotes", null, "mRNA was extracted from zygotes or by RIP experiments followed by Trizol extraction The RNA was extracted using chloroform and precipitated with iso propanol. Ovation RNA seq System V2 NuGEN library preperation kit was used and the library was sequenced on an Illumina HISEQ using 50bp single end sequencing.", "Zebrafish were maintained under standard conditions.", "fraction:Tdrd6a IP", "GSM2087138", "GSM2087138: Tdrd6a IP 3; Danio rerio; RIP Seq", "GSM2087138", null, "1", "mRNA was extracted from zygotes or by RIP experiments followed by Trizol extraction The RNA was extracted using chloroform and precipitated with iso propanol. Ovation RNA seq System V2 NuGEN library preperation kit was used and the library was sequenced on an Illumina HISEQ using 50bp single end sequencing.", "GEO Accession:GSM2087138", "RIP-Seq", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2000", null, "SRP071849", null, null, "WT_Tdrd6a_IP_3.fastq.gz", "fastq", 843641439.0, 16541989.0, "GSM2087138 r1", "0:51", "A:216465581;C:196892096;G:207722567;T:222361382;N:199813", 51, null, null, null, 216465581, 196892096, 207722567, 222361382, 199813, "SRX1637108", "SRS1342929", "SRA385813", "GEO", "Rene Ketting, RNA silencing, IMB", 1, 0.84615, null, 0.08957, null, 0.78768, null, 0.50612, null, 51, null, "B", null, "usable mapping rate", "illumina", "hiseq_era", "unknown", "other", "unknown", "sc_generic", "single_cell_generic", "generic-scrnaseq-only", null, "Germany", "2016-03-13", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [40705, "SRR3231359", "SRX1637107", "SRS1342930", "SRP071849", "PRJNA315400", "Tdrd6a recruits Ziwi bound piRNAs and coordinates deposition of germ plasm mRNAs into primordial germ cells [RIP Seq]", "GSE79161", "Transcriptome Analysis", "Germ plasm the Balbiani body and nuage are evolutionary conserved structures essential for germ cell specification and maintenance. We describe Tdrd6a as a component of these structures with two distinct molecular functions. First Tdrd6a facilitates the accumulation of the typical antisense bias of piRNAs without xxx effects on piRNA biogenesis signatures. Second we show that Tdrd6a is required for Balbiani body and germ plasm integrity and associates with RNA binding proteins and germ plasm mRNAs. On the cell biological level maternally contributed Tdrd6a strongly impacts germ cell formation but is dispensable for fertility. Using single cell RNA sequencing we demonstrate that Tdrd6a promotes early germ cell development and regulates the stoichiometry of germ plasm mRNAs. We propose that Tdrd6a functions as a scaffold to recruit correct ratios of germ plasm transcripts and to accumulate antisense piRNA complexes in order to ensure both specification and maintenance of germ cells. Overall design: RNA was extracted from ovary tissue or from RIP experiments by Trizol extraction.", "parent bioproject:PRJNA315403", "pubmed:30086300", null, "Tdrd6a IP 2", "GSM2087137", null, "tissue:Zebrafish Zygotes|fraction:Tdrd6a IP", "Tdrd6a IP 2", "Reads were mapped to Zv9 using tophat trapnell et al 2009to the ENSEMBLE gene build and further processing of the read counts was performed using Deseq Anders et al 2009 Genome build: zv9 Supplementary files format and content: The mRNA counts.csv contains the DESeq normalized read counts per Ensemble gene as indicated. Headers indicate the sample name.", "Zebrafish Zygotes", null, "mRNA was extracted from zygotes or by RIP experiments followed by Trizol extraction The RNA was extracted using chloroform and precipitated with iso propanol. Ovation RNA seq System V2 NuGEN library preperation kit was used and the library was sequenced on an Illumina HISEQ using 50bp single end sequencing.", "Zebrafish were maintained under standard conditions.", "fraction:Tdrd6a IP", "GSM2087137", "GSM2087137: Tdrd6a IP 2; Danio rerio; RIP Seq", "GSM2087137", null, "1", "mRNA was extracted from zygotes or by RIP experiments followed by Trizol extraction The RNA was extracted using chloroform and precipitated with iso propanol. Ovation RNA seq System V2 NuGEN library preperation kit was used and the library was sequenced on an Illumina HISEQ using 50bp single end sequencing.", "GEO Accession:GSM2087137", "RIP-Seq", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2000", null, "SRP071849", null, null, "WT_Tdrd6a_IP_2.fastq.gz", "fastq", 391198050.0, 7670550.0, "GSM2087137 r1", "0:51", "A:99015594;C:91525929;G:97155710;T:103371571;N:129246", 51, null, null, null, 99015594, 91525929, 97155710, 103371571, 129246, "SRX1637107", "SRS1342930", "SRA385813", "GEO", "Rene Ketting, RNA silencing, IMB", 1, 0.85778, null, 0.12015, null, 0.80515, null, 0.47541, null, 51, null, "B", null, "usable mapping rate", "illumina", "hiseq_era", "unknown", "other", "unknown", "sc_generic", "single_cell_generic", "generic-scrnaseq-only", null, "Germany", "2016-03-13", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [40706, "SRR3231358", "SRX1637106", "SRS1342931", "SRP071849", "PRJNA315400", "Tdrd6a recruits Ziwi bound piRNAs and coordinates deposition of germ plasm mRNAs into primordial germ cells [RIP Seq]", "GSE79161", "Transcriptome Analysis", "Germ plasm the Balbiani body and nuage are evolutionary conserved structures essential for germ cell specification and maintenance. We describe Tdrd6a as a component of these structures with two distinct molecular functions. First Tdrd6a facilitates the accumulation of the typical antisense bias of piRNAs without xxx effects on piRNA biogenesis signatures. Second we show that Tdrd6a is required for Balbiani body and germ plasm integrity and associates with RNA binding proteins and germ plasm mRNAs. On the cell biological level maternally contributed Tdrd6a strongly impacts germ cell formation but is dispensable for fertility. Using single cell RNA sequencing we demonstrate that Tdrd6a promotes early germ cell development and regulates the stoichiometry of germ plasm mRNAs. We propose that Tdrd6a functions as a scaffold to recruit correct ratios of germ plasm transcripts and to accumulate antisense piRNA complexes in order to ensure both specification and maintenance of germ cells. Overall design: RNA was extracted from ovary tissue or from RIP experiments by Trizol extraction.", "parent bioproject:PRJNA315403", "pubmed:30086300", null, "Tdrd6a IP 1", "GSM2087136", null, "tissue:Zebrafish Zygotes|fraction:Tdrd6a IP", "Tdrd6a IP 1", "Reads were mapped to Zv9 using tophat trapnell et al 2009to the ENSEMBLE gene build and further processing of the read counts was performed using Deseq Anders et al 2009 Genome build: zv9 Supplementary files format and content: The mRNA counts.csv contains the DESeq normalized read counts per Ensemble gene as indicated. Headers indicate the sample name.", "Zebrafish Zygotes", null, "mRNA was extracted from zygotes or by RIP experiments followed by Trizol extraction The RNA was extracted using chloroform and precipitated with iso propanol. Ovation RNA seq System V2 NuGEN library preperation kit was used and the library was sequenced on an Illumina HISEQ using 50bp single end sequencing.", "Zebrafish were maintained under standard conditions.", "fraction:Tdrd6a IP", "GSM2087136", "GSM2087136: Tdrd6a IP 1; Danio rerio; RIP Seq", "GSM2087136", null, "1", "mRNA was extracted from zygotes or by RIP experiments followed by Trizol extraction The RNA was extracted using chloroform and precipitated with iso propanol. Ovation RNA seq System V2 NuGEN library preperation kit was used and the library was sequenced on an Illumina HISEQ using 50bp single end sequencing.", "GEO Accession:GSM2087136", "RIP-Seq", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2000", null, "SRP071849", null, null, "WT_Tdrd6a_IP_1.fastq.gz", "fastq", 1927415052.0, 37792452.0, "GSM2087136 r1", "0:51", "A:498125298;C:444860381;G:468826607;T:515142356;N:460410", 51, null, null, null, 498125298, 444860381, 468826607, 515142356, 460410, "SRX1637106", "SRS1342931", "SRA385813", "GEO", "Rene Ketting, RNA silencing, IMB", 1, 0.86194, null, 0.09753, null, 0.80146, null, 0.47808, null, 51, null, "B", null, "usable mapping rate", "illumina", "hiseq_era", "unknown", "other", "unknown", "sc_generic", "single_cell_generic", "generic-scrnaseq-only", null, "Germany", "2016-03-13", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [41560, "SRR5017063", "SRX2345558", "SRS1796155", "SRP093295", "PRJNA353372", "N6 methyladenosine dynamics during early vertebrate embryogenesis", "GSE89815", "Other", "Early vertebrate embryogenesis is characterized by extensive post transcriptional regulation during the maternal to zygotic transition. The N6 methyladenosine m6A modifications on mRNA has been shown to affect both translation and stability of transcripts. Here we investigate the m6A topology during early vertebrate embryogenesis and its association with RNA stability translation efficiency and effect on miR 430 degradation kinetics. Notably we find a strong association of m6A with cytoplasmic polyadenylation and translational efficiency prior to zygotic genome activation. Genes required for zygotic genome activation such as nanog and pou5f3 display dynamic m6A levels. post zygotic genome activation m6A is associated with improved stability and dampens the effect of miR 430 mediated degradation. Through sequence analyses we identified enrichment of motifs for RNA binding proteins involved in translational regulation and RNA degradation. We propose a role for m6A in multiple mRNA regulatory mechanisms for the first time in an in vivo system and improve our understanding of the combinatorial code behind the complex post transcriptional regulation of reprogramming during early vertebrate development. Overall design: Examination of m6A in four different developmental stages", null, null, null, "input 1cell rep2", "GSM2390016", null, "tissue:Embryos|developmental stage:1cell embryos|strain:AB wild type", "input 1cell rep2", "Base calling The sequencing output from each of the 4 lanes the sequencing run was trimmed 7 nucleotides in the 5\u2019end and de multiplexed 4 samples in each lane. Reads were mapped to Zv10 using the STAR aligner Dobin et al. 2013 with options seedSearchStartLmax 15 clip3pNbases 10 clip5pNbases 10 outFilterMultimapNmax 20 outFilterMismatchNoverLmax 0.05 outFilterMatchNminOverLread 0.0 outFilterMatchNmin 15 outFilterScoreMinOverLread 0.0. We performed peak calling and detection of differentially methylated genes using ExomePeak Meng et al. 2013. Genome build: Zv10 Supplementary files format and content: Bed files with enriched regions from each developmental stage and txt file with raw and normalized read counts for each gene using the input samples", "Embryos", null, "We isolated total RNA from 4 stages 1 cell/20 minutes post fertilization 2 4 and 6 hpf in batches of 200 embryos using TRIzol Invitrogen cat.no. 15596 018. We added ERCC spike in RNA TermoFisher scientific cat.no. 4456740 to the trizol. For each sequencing library two batches of total RNA from 200 embryos were merged and enriched for polyA+ RNA using Dynabeads\u00ae mRNA Purification Kit Ambion #61006. The m6A RIP experiment was carried out as previously described Ke et al. 2015 and the protocol can be found in supplementary file 1. Briefly polyA+ enriched RNA was partially fragmented by alkaline hydrolysis ethanol precipitated and SDS PAGE size selected for 20 80nt. Part of the fragmented RNA was used as input and the rest was immunoprecipitated at 4oC for 2 h using Dynabeads Protein A Life Technologies # 10008D conjugated anti m6A antibody Synaptic systems # 202003. post stringent washing the bound RNA was eluted with 0.5mg/ml N6 methyladenosine sodium salt Sigma Aldrich # M2780 ethanol precipitated and resuspended with RNase free water. The eluted RNA and input RNA were subjected to 3\u2019pre adenylated DNA liker ligation with T4 RNA ligase2 truncated KQ NEB #M0373L at 16 oC over night. The sequencing library was constructed with bromodeoxyuridine BrdU CLIP protocol described in Weyn Vanhentenryck et al. 2014 with improved RT primers indicated in Ke et al. 2015.", "Embryos from the AB wild type strain were obtained from the NMBU zebrafish facility where the zebrafish were kept at 28\u00b11\u00b0C on a 14 10 hour light dark cycle at a density of 5 10 fish/L. System water SW was prepared from particle and active charcoal filtrated tap water deionized by reverse osmosis RO and kept sterile by UV irradiation. The water was conditioned to a conductivity of 500\u00b5S/cm general hardness GH 4 5 and pH 7.5 by addition of 155g synthetic sea salt Instant Ocean Blacksburg USA 53g sodium carbonate and 15g calcium chloride Sigma Aldrich per liter RO water. Adult fish were fed with Gemma Micro 300 Skretting Stavanger Norway dry feed twice a day and live artemia Scanbur Karlslunde Denmark once a day. Health monitoring was by daily inspection use of sentinel fish sent for pathology ZIRC Eugene Oregon and water microbiology analysis NMBU Vetbio Oslo every six month. Adult fish were allowed to mate for 30 minutes in standard 1L breeding tanks Aquatic Habitats Apopka FL. Harvested embryos were kept in autoclaved SW at 28 \u00b0C harvested by snap freezing liquid nitrogen and visually controlled for stage and lack of abnormalities at the selected time points. All experiments were performed according to Norwegian Animal Welfare Act 2009 the EU Directive 2010/63.", "developmental stage:1cell embryos|strain:AB wild type", "GSM2390016", "GSM2390016: input 1cell rep2; Danio rerio; RIP Seq", "GSM2390016", null, "1", "We isolated total RNA from 4 stages 1 cell/20 minutes post fertilization 2 4 and 6 hpf in batches of 200 embryos using TRIzol Invitrogen cat.no. 15596 018. We added ERCC spike in RNA TermoFisher scientific cat.no. 4456740 to the trizol. For each sequencing library two batches of total RNA from 200 embryos were merged and enriched for polyA+ RNA using Dynabeads\u00ae mRNA Purification Kit Ambion #61006. The m6A RIP experiment was carried out as previously described Ke et al. 2015 and the protocol can be found in supplementary file 1. Briefly polyA+ enriched RNA was partially fragmented by alkaline hydrolysis ethanol precipitated and SDS PAGE size selected for 20 80nt. Part of the fragmented RNA was used as input and the rest was immunoprecipitated at 4oC for 2 h using Dynabeads Protein A Life Technologies # 10008D conjugated anti m6A antibody Synaptic systems # 202003. post stringent washing the bound RNA was eluted with 0.5mg/ml N6 methyladenosine sodium salt Sigma Aldrich # M2780 ethanol precipitated and resuspended with RNase free water. The eluted RNA and input RNA were subjected to 3\u2019pre adenylated DNA liker ligation with T4 RNA ligase2 truncated KQ NEB #M0373L at 16 oC over night. The sequencing library was constructed with bromodeoxyuridine BrdU CLIP protocol described in Weyn Vanhentenryck et al. 2014 with improved RT primers indicated in Ke et al. 2015.", "GEO Accession:GSM2390016", "RIP-Seq", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2000", null, "SRP093295", null, null, "input_1cell_rep2.fastq.gz", "fastq", 3379762010.0, 35954915.0, "GSM2390016 r1", "0:94", "A:898920912;C:816234808;G:823428476;T:840956466;N:221348", 94, null, null, null, 898920912, 816234808, 823428476, 840956466, 221348, "SRX2345558", "SRS1796155", "SRA492943", "GEO", "Klungland Lab, Dept of microbiology, Oslo University Hospital", 1, 0.04513, null, 0.00864, null, 0.95085, null, 0.51692, null, 94, null, "B", null, "usable mapping rate", "illumina", "hiseq_era", "unknown", "poly_a", "unknown", "bulk", "unknown", "unknown", null, "Norway", "2016-11-14", "Zygote", "Embryo", "Embryo Imprecise", "All anatomical structures"], [41561, "SRR5017062", "SRX2345557", "SRS1796128", "SRP093295", "PRJNA353372", "N6 methyladenosine dynamics during early vertebrate embryogenesis", "GSE89815", "Other", "Early vertebrate embryogenesis is characterized by extensive post transcriptional regulation during the maternal to zygotic transition. The N6 methyladenosine m6A modifications on mRNA has been shown to affect both translation and stability of transcripts. Here we investigate the m6A topology during early vertebrate embryogenesis and its association with RNA stability translation efficiency and effect on miR 430 degradation kinetics. Notably we find a strong association of m6A with cytoplasmic polyadenylation and translational efficiency prior to zygotic genome activation. Genes required for zygotic genome activation such as nanog and pou5f3 display dynamic m6A levels. post zygotic genome activation m6A is associated with improved stability and dampens the effect of miR 430 mediated degradation. Through sequence analyses we identified enrichment of motifs for RNA binding proteins involved in translational regulation and RNA degradation. We propose a role for m6A in multiple mRNA regulatory mechanisms for the first time in an in vivo system and improve our understanding of the combinatorial code behind the complex post transcriptional regulation of reprogramming during early vertebrate development. Overall design: Examination of m6A in four different developmental stages", null, null, null, "input 1cell rep1", "GSM2390015", null, "tissue:Embryos|developmental stage:1cell embryos|strain:AB wild type", "input 1cell rep1", "Base calling The sequencing output from each of the 4 lanes the sequencing run was trimmed 7 nucleotides in the 5\u2019end and de multiplexed 4 samples in each lane. Reads were mapped to Zv10 using the STAR aligner Dobin et al. 2013 with options seedSearchStartLmax 15 clip3pNbases 10 clip5pNbases 10 outFilterMultimapNmax 20 outFilterMismatchNoverLmax 0.05 outFilterMatchNminOverLread 0.0 outFilterMatchNmin 15 outFilterScoreMinOverLread 0.0. We performed peak calling and detection of differentially methylated genes using ExomePeak Meng et al. 2013. Genome build: Zv10 Supplementary files format and content: Bed files with enriched regions from each developmental stage and txt file with raw and normalized read counts for each gene using the input samples", "Embryos", null, "We isolated total RNA from 4 stages 1 cell/20 minutes post fertilization 2 4 and 6 hpf in batches of 200 embryos using TRIzol Invitrogen cat.no. 15596 018. We added ERCC spike in RNA TermoFisher scientific cat.no. 4456740 to the trizol. For each sequencing library two batches of total RNA from 200 embryos were merged and enriched for polyA+ RNA using Dynabeads\u00ae mRNA Purification Kit Ambion #61006. The m6A RIP experiment was carried out as previously described Ke et al. 2015 and the protocol can be found in supplementary file 1. Briefly polyA+ enriched RNA was partially fragmented by alkaline hydrolysis ethanol precipitated and SDS PAGE size selected for 20 80nt. Part of the fragmented RNA was used as input and the rest was immunoprecipitated at 4oC for 2 h using Dynabeads Protein A Life Technologies # 10008D conjugated anti m6A antibody Synaptic systems # 202003. post stringent washing the bound RNA was eluted with 0.5mg/ml N6 methyladenosine sodium salt Sigma Aldrich # M2780 ethanol precipitated and resuspended with RNase free water. The eluted RNA and input RNA were subjected to 3\u2019pre adenylated DNA liker ligation with T4 RNA ligase2 truncated KQ NEB #M0373L at 16 oC over night. The sequencing library was constructed with bromodeoxyuridine BrdU CLIP protocol described in Weyn Vanhentenryck et al. 2014 with improved RT primers indicated in Ke et al. 2015.", "Embryos from the AB wild type strain were obtained from the NMBU zebrafish facility where the zebrafish were kept at 28\u00b11\u00b0C on a 14 10 hour light dark cycle at a density of 5 10 fish/L. System water SW was prepared from particle and active charcoal filtrated tap water deionized by reverse osmosis RO and kept sterile by UV irradiation. The water was conditioned to a conductivity of 500\u00b5S/cm general hardness GH 4 5 and pH 7.5 by addition of 155g synthetic sea salt Instant Ocean Blacksburg USA 53g sodium carbonate and 15g calcium chloride Sigma Aldrich per liter RO water. Adult fish were fed with Gemma Micro 300 Skretting Stavanger Norway dry feed twice a day and live artemia Scanbur Karlslunde Denmark once a day. Health monitoring was by daily inspection use of sentinel fish sent for pathology ZIRC Eugene Oregon and water microbiology analysis NMBU Vetbio Oslo every six month. Adult fish were allowed to mate for 30 minutes in standard 1L breeding tanks Aquatic Habitats Apopka FL. Harvested embryos were kept in autoclaved SW at 28 \u00b0C harvested by snap freezing liquid nitrogen and visually controlled for stage and lack of abnormalities at the selected time points. All experiments were performed according to Norwegian Animal Welfare Act 2009 the EU Directive 2010/63.", "developmental stage:1cell embryos|strain:AB wild type", "GSM2390015", "GSM2390015: input 1cell rep1; Danio rerio; RIP Seq", "GSM2390015", null, "1", "We isolated total RNA from 4 stages 1 cell/20 minutes post fertilization 2 4 and 6 hpf in batches of 200 embryos using TRIzol Invitrogen cat.no. 15596 018. We added ERCC spike in RNA TermoFisher scientific cat.no. 4456740 to the trizol. For each sequencing library two batches of total RNA from 200 embryos were merged and enriched for polyA+ RNA using Dynabeads\u00ae mRNA Purification Kit Ambion #61006. The m6A RIP experiment was carried out as previously described Ke et al. 2015 and the protocol can be found in supplementary file 1. Briefly polyA+ enriched RNA was partially fragmented by alkaline hydrolysis ethanol precipitated and SDS PAGE size selected for 20 80nt. Part of the fragmented RNA was used as input and the rest was immunoprecipitated at 4oC for 2 h using Dynabeads Protein A Life Technologies # 10008D conjugated anti m6A antibody Synaptic systems # 202003. post stringent washing the bound RNA was eluted with 0.5mg/ml N6 methyladenosine sodium salt Sigma Aldrich # M2780 ethanol precipitated and resuspended with RNase free water. The eluted RNA and input RNA were subjected to 3\u2019pre adenylated DNA liker ligation with T4 RNA ligase2 truncated KQ NEB #M0373L at 16 oC over night. The sequencing library was constructed with bromodeoxyuridine BrdU CLIP protocol described in Weyn Vanhentenryck et al. 2014 with improved RT primers indicated in Ke et al. 2015.", "GEO Accession:GSM2390015", "RIP-Seq", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2000", null, "SRP093295", null, null, "input_1cell_rep1.fastq.gz", "fastq", 3650321646.0, 38833209.0, "GSM2390015 r1", "0:94", "A:951411355;C:878458528;G:890250453;T:929968044;N:233266", 94, null, null, null, 951411355, 878458528, 890250453, 929968044, 233266, "SRX2345557", "SRS1796128", "SRA492943", "GEO", "Klungland Lab, Dept of microbiology, Oslo University Hospital", 1, 0.03728, null, 0.00668, null, 0.94901, null, 0.60038, null, 94, null, "B", null, "usable mapping rate", "illumina", "hiseq_era", "unknown", "poly_a", "unknown", "bulk", "unknown", "unknown", null, "Norway", "2016-11-14", "Zygote", "Embryo", "Embryo Imprecise", "All anatomical structures"], [41562, "SRR5017061", "SRX2345556", "SRS1796127", "SRP093295", "PRJNA353372", "N6 methyladenosine dynamics during early vertebrate embryogenesis", "GSE89815", "Other", "Early vertebrate embryogenesis is characterized by extensive post transcriptional regulation during the maternal to zygotic transition. The N6 methyladenosine m6A modifications on mRNA has been shown to affect both translation and stability of transcripts. Here we investigate the m6A topology during early vertebrate embryogenesis and its association with RNA stability translation efficiency and effect on miR 430 degradation kinetics. Notably we find a strong association of m6A with cytoplasmic polyadenylation and translational efficiency prior to zygotic genome activation. Genes required for zygotic genome activation such as nanog and pou5f3 display dynamic m6A levels. post zygotic genome activation m6A is associated with improved stability and dampens the effect of miR 430 mediated degradation. Through sequence analyses we identified enrichment of motifs for RNA binding proteins involved in translational regulation and RNA degradation. We propose a role for m6A in multiple mRNA regulatory mechanisms for the first time in an in vivo system and improve our understanding of the combinatorial code behind the complex post transcriptional regulation of reprogramming during early vertebrate development. Overall design: Examination of m6A in four different developmental stages", null, null, null, "ip 1cell rep2", "GSM2390014", null, "tissue:Embryos|developmental stage:1cell embryos|strain:AB wild type", "ip 1cell rep2", "Base calling The sequencing output from each of the 4 lanes the sequencing run was trimmed 7 nucleotides in the 5\u2019end and de multiplexed 4 samples in each lane. Reads were mapped to Zv10 using the STAR aligner Dobin et al. 2013 with options seedSearchStartLmax 15 clip3pNbases 10 clip5pNbases 10 outFilterMultimapNmax 20 outFilterMismatchNoverLmax 0.05 outFilterMatchNminOverLread 0.0 outFilterMatchNmin 15 outFilterScoreMinOverLread 0.0. We performed peak calling and detection of differentially methylated genes using ExomePeak Meng et al. 2013. Genome build: Zv10 Supplementary files format and content: Bed files with enriched regions from each developmental stage and txt file with raw and normalized read counts for each gene using the input samples", "Embryos", null, "We isolated total RNA from 4 stages 1 cell/20 minutes post fertilization 2 4 and 6 hpf in batches of 200 embryos using TRIzol Invitrogen cat.no. 15596 018. We added ERCC spike in RNA TermoFisher scientific cat.no. 4456740 to the trizol. For each sequencing library two batches of total RNA from 200 embryos were merged and enriched for polyA+ RNA using Dynabeads\u00ae mRNA Purification Kit Ambion #61006. The m6A RIP experiment was carried out as previously described Ke et al. 2015 and the protocol can be found in supplementary file 1. Briefly polyA+ enriched RNA was partially fragmented by alkaline hydrolysis ethanol precipitated and SDS PAGE size selected for 20 80nt. Part of the fragmented RNA was used as input and the rest was immunoprecipitated at 4oC for 2 h using Dynabeads Protein A Life Technologies # 10008D conjugated anti m6A antibody Synaptic systems # 202003. post stringent washing the bound RNA was eluted with 0.5mg/ml N6 methyladenosine sodium salt Sigma Aldrich # M2780 ethanol precipitated and resuspended with RNase free water. The eluted RNA and input RNA were subjected to 3\u2019pre adenylated DNA liker ligation with T4 RNA ligase2 truncated KQ NEB #M0373L at 16 oC over night. The sequencing library was constructed with bromodeoxyuridine BrdU CLIP protocol described in Weyn Vanhentenryck et al. 2014 with improved RT primers indicated in Ke et al. 2015.", "Embryos from the AB wild type strain were obtained from the NMBU zebrafish facility where the zebrafish were kept at 28\u00b11\u00b0C on a 14 10 hour light dark cycle at a density of 5 10 fish/L. System water SW was prepared from particle and active charcoal filtrated tap water deionized by reverse osmosis RO and kept sterile by UV irradiation. The water was conditioned to a conductivity of 500\u00b5S/cm general hardness GH 4 5 and pH 7.5 by addition of 155g synthetic sea salt Instant Ocean Blacksburg USA 53g sodium carbonate and 15g calcium chloride Sigma Aldrich per liter RO water. Adult fish were fed with Gemma Micro 300 Skretting Stavanger Norway dry feed twice a day and live artemia Scanbur Karlslunde Denmark once a day. Health monitoring was by daily inspection use of sentinel fish sent for pathology ZIRC Eugene Oregon and water microbiology analysis NMBU Vetbio Oslo every six month. Adult fish were allowed to mate for 30 minutes in standard 1L breeding tanks Aquatic Habitats Apopka FL. Harvested embryos were kept in autoclaved SW at 28 \u00b0C harvested by snap freezing liquid nitrogen and visually controlled for stage and lack of abnormalities at the selected time points. All experiments were performed according to Norwegian Animal Welfare Act 2009 the EU Directive 2010/63.", "developmental stage:1cell embryos|strain:AB wild type", "GSM2390014", "GSM2390014: ip 1cell rep2; Danio rerio; RIP Seq", "GSM2390014", null, "1", "We isolated total RNA from 4 stages 1 cell/20 minutes post fertilization 2 4 and 6 hpf in batches of 200 embryos using TRIzol Invitrogen cat.no. 15596 018. We added ERCC spike in RNA TermoFisher scientific cat.no. 4456740 to the trizol. For each sequencing library two batches of total RNA from 200 embryos were merged and enriched for polyA+ RNA using Dynabeads\u00ae mRNA Purification Kit Ambion #61006. The m6A RIP experiment was carried out as previously described Ke et al. 2015 and the protocol can be found in supplementary file 1. Briefly polyA+ enriched RNA was partially fragmented by alkaline hydrolysis ethanol precipitated and SDS PAGE size selected for 20 80nt. Part of the fragmented RNA was used as input and the rest was immunoprecipitated at 4oC for 2 h using Dynabeads Protein A Life Technologies # 10008D conjugated anti m6A antibody Synaptic systems # 202003. post stringent washing the bound RNA was eluted with 0.5mg/ml N6 methyladenosine sodium salt Sigma Aldrich # M2780 ethanol precipitated and resuspended with RNase free water. The eluted RNA and input RNA were subjected to 3\u2019pre adenylated DNA liker ligation with T4 RNA ligase2 truncated KQ NEB #M0373L at 16 oC over night. The sequencing library was constructed with bromodeoxyuridine BrdU CLIP protocol described in Weyn Vanhentenryck et al. 2014 with improved RT primers indicated in Ke et al. 2015.", "GEO Accession:GSM2390014", "RIP-Seq", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2000", null, "SRP093295", null, null, "ip_1cell_rep2.fastq.gz", "fastq", 3485432580.0, 37079070.0, "GSM2390014 r1", "0:94", "A:953699047;C:835179152;G:846963064;T:848487912;N:1103405", 94, null, null, null, 953699047, 835179152, 846963064, 848487912, 1103405, "SRX2345556", "SRS1796127", "SRA492943", "GEO", "Klungland Lab, Dept of microbiology, Oslo University Hospital", 1, 0.02534, null, 0.00219, null, 0.95891, null, 0.5975, null, 94, null, "B", null, "usable mapping rate", "illumina", "hiseq_era", "unknown", "poly_a", "unknown", "bulk", "unknown", "unknown", null, "Norway", "2016-11-14", "Zygote", "Embryo", "Embryo Imprecise", "All anatomical structures"], [41563, "SRR5017060", "SRX2345555", "SRS1796130", "SRP093295", "PRJNA353372", "N6 methyladenosine dynamics during early vertebrate embryogenesis", "GSE89815", "Other", "Early vertebrate embryogenesis is characterized by extensive post transcriptional regulation during the maternal to zygotic transition. The N6 methyladenosine m6A modifications on mRNA has been shown to affect both translation and stability of transcripts. Here we investigate the m6A topology during early vertebrate embryogenesis and its association with RNA stability translation efficiency and effect on miR 430 degradation kinetics. Notably we find a strong association of m6A with cytoplasmic polyadenylation and translational efficiency prior to zygotic genome activation. Genes required for zygotic genome activation such as nanog and pou5f3 display dynamic m6A levels. post zygotic genome activation m6A is associated with improved stability and dampens the effect of miR 430 mediated degradation. Through sequence analyses we identified enrichment of motifs for RNA binding proteins involved in translational regulation and RNA degradation. We propose a role for m6A in multiple mRNA regulatory mechanisms for the first time in an in vivo system and improve our understanding of the combinatorial code behind the complex post transcriptional regulation of reprogramming during early vertebrate development. Overall design: Examination of m6A in four different developmental stages", null, null, null, "ip 1cell rep1", "GSM2390013", null, "tissue:Embryos|developmental stage:1cell embryos|strain:AB wild type", "ip 1cell rep1", "Base calling The sequencing output from each of the 4 lanes the sequencing run was trimmed 7 nucleotides in the 5\u2019end and de multiplexed 4 samples in each lane. Reads were mapped to Zv10 using the STAR aligner Dobin et al. 2013 with options seedSearchStartLmax 15 clip3pNbases 10 clip5pNbases 10 outFilterMultimapNmax 20 outFilterMismatchNoverLmax 0.05 outFilterMatchNminOverLread 0.0 outFilterMatchNmin 15 outFilterScoreMinOverLread 0.0. We performed peak calling and detection of differentially methylated genes using ExomePeak Meng et al. 2013. Genome build: Zv10 Supplementary files format and content: Bed files with enriched regions from each developmental stage and txt file with raw and normalized read counts for each gene using the input samples", "Embryos", null, "We isolated total RNA from 4 stages 1 cell/20 minutes post fertilization 2 4 and 6 hpf in batches of 200 embryos using TRIzol Invitrogen cat.no. 15596 018. We added ERCC spike in RNA TermoFisher scientific cat.no. 4456740 to the trizol. For each sequencing library two batches of total RNA from 200 embryos were merged and enriched for polyA+ RNA using Dynabeads\u00ae mRNA Purification Kit Ambion #61006. The m6A RIP experiment was carried out as previously described Ke et al. 2015 and the protocol can be found in supplementary file 1. Briefly polyA+ enriched RNA was partially fragmented by alkaline hydrolysis ethanol precipitated and SDS PAGE size selected for 20 80nt. Part of the fragmented RNA was used as input and the rest was immunoprecipitated at 4oC for 2 h using Dynabeads Protein A Life Technologies # 10008D conjugated anti m6A antibody Synaptic systems # 202003. post stringent washing the bound RNA was eluted with 0.5mg/ml N6 methyladenosine sodium salt Sigma Aldrich # M2780 ethanol precipitated and resuspended with RNase free water. The eluted RNA and input RNA were subjected to 3\u2019pre adenylated DNA liker ligation with T4 RNA ligase2 truncated KQ NEB #M0373L at 16 oC over night. The sequencing library was constructed with bromodeoxyuridine BrdU CLIP protocol described in Weyn Vanhentenryck et al. 2014 with improved RT primers indicated in Ke et al. 2015.", "Embryos from the AB wild type strain were obtained from the NMBU zebrafish facility where the zebrafish were kept at 28\u00b11\u00b0C on a 14 10 hour light dark cycle at a density of 5 10 fish/L. System water SW was prepared from particle and active charcoal filtrated tap water deionized by reverse osmosis RO and kept sterile by UV irradiation. The water was conditioned to a conductivity of 500\u00b5S/cm general hardness GH 4 5 and pH 7.5 by addition of 155g synthetic sea salt Instant Ocean Blacksburg USA 53g sodium carbonate and 15g calcium chloride Sigma Aldrich per liter RO water. Adult fish were fed with Gemma Micro 300 Skretting Stavanger Norway dry feed twice a day and live artemia Scanbur Karlslunde Denmark once a day. Health monitoring was by daily inspection use of sentinel fish sent for pathology ZIRC Eugene Oregon and water microbiology analysis NMBU Vetbio Oslo every six month. Adult fish were allowed to mate for 30 minutes in standard 1L breeding tanks Aquatic Habitats Apopka FL. Harvested embryos were kept in autoclaved SW at 28 \u00b0C harvested by snap freezing liquid nitrogen and visually controlled for stage and lack of abnormalities at the selected time points. All experiments were performed according to Norwegian Animal Welfare Act 2009 the EU Directive 2010/63.", "developmental stage:1cell embryos|strain:AB wild type", "GSM2390013", "GSM2390013: ip 1cell rep1; Danio rerio; RIP Seq", "GSM2390013", null, "1", "We isolated total RNA from 4 stages 1 cell/20 minutes post fertilization 2 4 and 6 hpf in batches of 200 embryos using TRIzol Invitrogen cat.no. 15596 018. We added ERCC spike in RNA TermoFisher scientific cat.no. 4456740 to the trizol. For each sequencing library two batches of total RNA from 200 embryos were merged and enriched for polyA+ RNA using Dynabeads\u00ae mRNA Purification Kit Ambion #61006. The m6A RIP experiment was carried out as previously described Ke et al. 2015 and the protocol can be found in supplementary file 1. Briefly polyA+ enriched RNA was partially fragmented by alkaline hydrolysis ethanol precipitated and SDS PAGE size selected for 20 80nt. Part of the fragmented RNA was used as input and the rest was immunoprecipitated at 4oC for 2 h using Dynabeads Protein A Life Technologies # 10008D conjugated anti m6A antibody Synaptic systems # 202003. post stringent washing the bound RNA was eluted with 0.5mg/ml N6 methyladenosine sodium salt Sigma Aldrich # M2780 ethanol precipitated and resuspended with RNase free water. The eluted RNA and input RNA were subjected to 3\u2019pre adenylated DNA liker ligation with T4 RNA ligase2 truncated KQ NEB #M0373L at 16 oC over night. The sequencing library was constructed with bromodeoxyuridine BrdU CLIP protocol described in Weyn Vanhentenryck et al. 2014 with improved RT primers indicated in Ke et al. 2015.", "GEO Accession:GSM2390013", "RIP-Seq", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2000", null, "SRP093295", null, null, "ip_1cell_rep1.fastq.gz", "fastq", 3610167102.0, 38406033.0, "GSM2390013 r1", "0:94", "A:961438376;C:875781200;G:891055201;T:880749228;N:1143097", 94, null, null, null, 961438376, 875781200, 891055201, 880749228, 1143097, "SRX2345555", "SRS1796130", "SRA492943", "GEO", "Klungland Lab, Dept of microbiology, Oslo University Hospital", 1, 0.01939, null, 0.00159, null, 0.96457, null, 0.53652, null, 94, null, "B", null, "usable mapping rate", "illumina", "hiseq_era", "unknown", "poly_a", "unknown", "bulk", "unknown", "unknown", null, "Norway", "2016-11-14", "Zygote", "Embryo", "Embryo Imprecise", "All anatomical structures"], [44659, "SRR6268189", "SRX3374357", "SRS2671584", "SRP124609", "PRJNA417597", "Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation", "GSE106677", "Other", "The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were collected at 1h 2h 3h 4h 3 samples 6h 8h and 10h.", null, "pubmed:29225039", null, "rep A uninjected", "GSM2845343", null, "source name:zebrafish embryos|developmental stage:NA|tissue:embryo", "rep A uninjected", "Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5\u2019 terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels", "zebrafish embryos", "Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage.", "Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3\u2019UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3\u2019UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3\u2019UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors", "Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage.", "developmental stage:NA|tissue:embryo", "GSM2845343", "GSM2845343: rep A uninjected; Danio rerio; OTHER", "GSM2845343", null, "1", "Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors", "GEO Accession:GSM2845343", "OTHER", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina MiSeq", null, "SRP124609", null, null, "SpeI-pool.fastq.gz", "fastq", 2026742240.0, 12667139.0, "GSM2845343 r1", "0:160 1:0", "A:625744065;C:476430927;G:385335893;T:539051959;N:179396", 160, 0, null, null, 625744065, 476430927, 385335893, 539051959, 179396, "SRX3374357", "SRS2671584", "SRA629220", "GEO", "Broad Institute", 1, 4e-05, null, 0.0, null, 0.99987, null, 0.16666, null, 160, null, "T", null, "under 1.2% mapping rate", "illumina", "miseq", "unknown", "random_priming", "unknown", "bulk", "unknown", "unknown", null, "United States", "2017-11-08", "Zygote", "Embryo", "Embryo Imprecise", "All anatomical structures"], [44671, "SRR6268177", "SRX3374345", "SRS2671574", "SRP124609", "PRJNA417597", "Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation", "GSE106677", "Other", "The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were collected at 1h 2h 3h 4h 3 samples 6h 8h and 10h.", null, "pubmed:29225039", null, "techrep A+ uninjected.2", "GSM2845331", null, "source name:zebrafish embryos|developmental stage:NA|tissue:embryo", "techrep A+ uninjected.2", "Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5\u2019 terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels", "zebrafish embryos", "Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage.", "Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3\u2019UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3\u2019UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3\u2019UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors", "Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage.", "developmental stage:NA|tissue:embryo", "GSM2845331", "GSM2845331: techrep A+ uninjected.2; Danio rerio; OTHER", "GSM2845331", null, "1", "Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors", "GEO Accession:GSM2845331", "OTHER", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina MiSeq", null, "SRP124609", null, null, "AR13_S13.fastq.gz", "fastq", 206761464.0, 1230723.0, "GSM2845331 r1", "0:168 1:0", "A:70709853;C:42141255;G:36769712;T:57139973;N:671", 168, 0, null, null, 70709853, 42141255, 36769712, 57139973, 671, "SRX3374345", "SRS2671574", "SRA629220", "GEO", "Broad Institute", 1, 1e-05, null, 0.0, null, 1.0, null, null, null, 168, null, "T", null, "under 1.2% mapping rate", "illumina", "miseq", "unknown", "random_priming", "unknown", "bulk", "unknown", "unknown", null, "United States", "2017-11-08", "Zygote", "Embryo", "Embryo Imprecise", "All anatomical structures"], [44683, "SRR6268165", "SRX3374333", "SRS2671560", "SRP124609", "PRJNA417597", "Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation", "GSE106677", "Other", "The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were collected at 1h 2h 3h 4h 3 samples 6h 8h and 10h.", null, "pubmed:29225039", null, "techrep A+ uninjected.1", "GSM2845319", null, "source name:zebrafish embryos|developmental stage:NA|tissue:embryo", "techrep A+ uninjected.1", "Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5\u2019 terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels", "zebrafish embryos", "Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage.", "Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3\u2019UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3\u2019UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3\u2019UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors", "Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage.", "developmental stage:NA|tissue:embryo", "GSM2845319", "GSM2845319: techrep A+ uninjected.1; Danio rerio; OTHER", "GSM2845319", null, "1", "Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors", "GEO Accession:GSM2845319", "OTHER", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina MiSeq", null, "SRP124609", null, null, "AR01_S1.fastq.gz", "fastq", 298586232.0, 1777299.0, "GSM2845319 r1", "0:168 1:0", "A:97160368;C:64341107;G:58562707;T:78521174;N:876", 168, 0, null, null, 97160368, 64341107, 58562707, 78521174, 876, "SRX3374333", "SRS2671560", "SRA629220", "GEO", "Broad Institute", 1, 1e-05, null, 0.0, null, 0.99997, null, 0.0, null, 168, null, "T", null, "under 1.2% mapping rate", "illumina", "miseq", "unknown", "random_priming", "unknown", "bulk", "unknown", "unknown", null, "United States", "2017-11-08", "Zygote", "Embryo", "Embryo Imprecise", "All anatomical structures"], [44694, "SRR6268154", "SRX3374322", "SRS2671551", "SRP124609", "PRJNA417597", "Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation", "GSE106677", "Other", "The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were collected at 1h 2h 3h 4h 3 samples 6h 8h and 10h.", null, "pubmed:29225039", null, "biorep A+ uninjected", "GSM2845308", null, "source name:zebrafish embryos|developmental stage:NA|tissue:embryo", "biorep A+ uninjected", "Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5\u2019 terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels", "zebrafish embryos", "Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage.", "Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3\u2019UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3\u2019UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3\u2019UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors", "Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage.", "developmental stage:NA|tissue:embryo", "GSM2845308", "GSM2845308: biorep A+ uninjected; Danio rerio; OTHER", "GSM2845308", null, "1", "Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors", "GEO Accession:GSM2845308", "OTHER", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina MiSeq", null, "SRP124609", null, null, "pool_S7.fastq.gz", "fastq", 818166560.0, 5113541.0, "GSM2845308 r1", "0:160 1:0", "A:256677558;C:187984574;G:154806708;T:218648792;N:48928", 160, 0, null, null, 256677558, 187984574, 154806708, 218648792, 48928, "SRX3374322", "SRS2671551", "SRA629220", "GEO", "Broad Institute", 1, 8e-05, null, 0.0, null, 0.99975, null, 0.5, null, 160, null, "T", null, "under 1.2% mapping rate", "illumina", "miseq", "unknown", "random_priming", "unknown", "bulk", "unknown", "unknown", null, "United States", "2017-11-08", "Zygote", "Embryo", "Embryo Imprecise", "All anatomical structures"], [44704, "SRR6268144", "SRX3374312", "SRS2671541", "SRP124609", "PRJNA417597", "Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation", "GSE106677", "Other", "The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were collected at 1h 2h 3h 4h 3 samples 6h 8h and 10h.", null, "pubmed:29225039", null, "embryo A uninjected", "GSM2845298", null, "source name:zebrafish embryos|developmental stage:NA|tissue:embryo", "embryo A uninjected", "Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5\u2019 terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels", "zebrafish embryos", "Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage.", "Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3\u2019UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3\u2019UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3\u2019UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors", "Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage.", "developmental stage:NA|tissue:embryo", "GSM2845298", "GSM2845298: embryo A uninjected; Danio rerio; OTHER", "GSM2845298", null, "1", "Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors", "GEO Accession:GSM2845298", "OTHER", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP124609", null, null, "SPE_0xA_AR01.fastq.gz", "fastq", 700784500.0, 7007845.0, "GSM2845298 r1", "0:100", "A:232295406;C:134437574;G:139352563;T:194535978;N:162979", 100, null, null, null, 232295406, 134437574, 139352563, 194535978, 162979, "SRX3374312", "SRS2671541", "SRA629220", "GEO", "Broad Institute", 1, 4e-05, null, 0.0, null, 0.99987, null, 0.16666, null, 100, null, "T", null, "under 1.2% mapping rate", "illumina", "hiseq_era", "unknown", "random_priming", "unknown", "bulk", "unknown", "unknown", null, "United States", "2017-11-08", "Zygote", "Embryo", "Embryo Imprecise", "All anatomical structures"], [44714, "SRR6268134", "SRX3374302", "SRS2671533", "SRP124609", "PRJNA417597", "Massively parallel reporter assay of three primeUTR sequences identifies in vivo rules for mRNA degradation", "GSE106677", "Other", "The stability of mRNAs is regulated by signals within their sequences but a systematic and predictive understanding of the underlying sequence rules remains elusive. Here we introduce UTR Seq a combination of massively parallel reporter assays and regression models to survey the dynamics of tens of thousands of three primeUTR sequences during early zebrafish embryogenesis. UTR Seq revealed two temporal degradation programs: a maternally encoded early onset program and a late onset program that accelerated degradation post zygotic genome activation. Three signals regulated early onset rates: stabilizing poly U and UUAG sequences and destabilizing GC rich signals. Three signals explained late onset degradation: miR 430 seeds AU rich sequences and Pumilio recognition sites. Sequence based regression models translated three primeUTRs into their unique decay patterns and predicted the in vivo impact of sequence signals on mRNA stability. Their application led to the successful design of artificial three primeUTRs that conferred specific mRNA dynamics. UTR Seq provides a general strategy to uncover the rules of RNA cis regulation. Overall design: temporal expression profiles of reporter mRNAs from zebrafish embryos 7 replicates total or oocytes 2 replicates total. A total of five embryo replicates were collected using pre adenylated A+ reporters in three separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment a single sample was collected every hour between 1h to 10h and split into two post RNA extraction to produce two technical replicates techrep. In the third experiment two separate samples were collected every two hours between 2h to 10h to produce two same day biological replicates biorep. A total of two replicates were collected using non adenylated A reporters in two separate experiments. In the first experiment a sample was collected every hour between 1h to 8h and at 10h. In the second experiment rep samples were collected at 1h 2h 3h 4h 3 samples 6h 8h and 10h.", null, "pubmed:29225039", null, "embryo A+ uninjected", "GSM2845288", null, "source name:zebrafish embryos|developmental stage:NA|tissue:embryo", "embryo A+ uninjected", "Library strategy: UTR Seq data 160nt reads was filtered to retain only sequences that contained both terminal adapter sequences with up to 10 mismatches and an insert of 90nt or longer. data 100nt reads was filtered to retain only sequences that contained the 5\u2019 terminal adapter sequence with up to 10 mismatches and an insert of 62nt or longer. Bowtie2 was used to align retained reads to a reference set of all 90 000 synthetic oligonucleotide sequences The number of UMIs that were mapped to each oligonucleotide sequence was recorded and adjusted to represent the expected number of mRNA molecules in the sample. A generalized binomial linear regression model was fitted to counts of five control mRNAs that were added to samples at known quantities in 2 fold increments highest 50fg lowest 3.125fg and the resulting linear transformation was used to normalize UMI counts. Genome build: The file sequences.txt is a FASTA file with the oligonucleotide sequences used for real alignment. This file is available on the series record. Supplementary files format and content: tab delimited files of normalized mRNA levels", "zebrafish embryos", "Embryos/oocytes were removed from their chorion and injected with 50 80pg of reporter mRNAs at the one cell stage.", "Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known 3\u2019UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant 3\u2019UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3\u2019UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors", "Fertilized zebrafish eggs or oocytes were collected at 28C and kept in culture medium 5.03mM NaCl 0.17mM KCl 0.33mM CaCl2 0.33mM MgSo4 0.1% Methylene blue. A total of 20 40 injected embryos/oocytes were randomly collected per sample post ensuring that all embryos were at the same expected developmental stage.", "developmental stage:NA|tissue:embryo", "GSM2845288", "GSM2845288: embryo A+ uninjected; Danio rerio; OTHER", "GSM2845288", null, "1", "Total RNA was isolated using TRIzol Invitrogen post adding 120fg of mRNA with 5 known three primeUTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step total RNA was reverse transcribed with Maxima RT Thermo Fisher and a gene specific primer that matched the constant three primeUTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant three primeUTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors", "GEO Accession:GSM2845288", "OTHER", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP124609", null, null, "SPE_40xA_AR01.fastq.gz", "fastq", 861781700.0, 8617817.0, "GSM2845288 r1", "0:100", "A:284066965;C:166650973;G:172491916;T:238392534;N:179312", 100, null, null, null, 284066965, 166650973, 172491916, 238392534, 179312, "SRX3374302", "SRS2671533", "SRA629220", "GEO", "Broad Institute", 1, 5e-05, null, 0.0, null, 0.99985, null, 0.125, null, 100, null, "T", null, "under 1.2% mapping rate", "illumina", "hiseq_era", "unknown", "random_priming", "unknown", "bulk", "unknown", "unknown", null, "United States", "2017-11-08", "Zygote", "Embryo", "Embryo Imprecise", "All anatomical structures"], [49538, "SRR8937006", "SRX5717519", "SRS4655939", "SRP162876", "PRJNA493828", "Ybx1 binding sites and Ybx1 binding m5C sites in zebrafish embryo sample.", "GSE120646", "Other", "The maternal to zygotic transition MZT is a conserved and fundamental process during which the embryo undergoes dramatic reprogramming to convert maternal environment to embryonic driven programing. However how the maternally supplied transcripts are dynamically regulated during MZT remains largely unknown. Herein through genome wide profiling of RNA 5 methylcytosine m5C in zebrafish early embryos we show that m5C methylated maternal mRNAs display higher stability during MZT. We identify that the Y box binding protein 1 Ybx1 prefers to recognizing m5C modified mRNAs through p p interaction with a key residue Trp45 in its cold shock domain CSD which plays essential roles in maternal mRNA stability and early embryogenesis of zebrafish. Cooperated with an mRNA stabilizer Pabpc1a Ybx1 promotes the stability of its target mRNAs in an m5C dependent manner. Our study demonstrates a novel mechanism of RNA m5C methylation regulated maternal mRNA stability during zebrafish MZT highlighting the critical role of m5C mRNA methylation in early development. Overall design: Examination of Ybx1 binding sites and Ybx1 binding m5C sites in zebrafish embryo. RIP seq: Briefly 500 zebrafish embryos at shield stage 4 hpf were resuspended with 2 ml lysis buffer 150 mM KCl 10 mM HEPES pH 7.6 2 mM EDTA 0.5% NP 40 0.5 mM DTT 1:100 protease inhibitor cocktail 0.4 U/\u00ce\u00bcl RNasin by rotating at 4\u00c2\u00b0C for 30 min and centrifuged at 15 000 g for 15 min. The supernatant was collected and pre cleared with protein A Dynabeads by rotation at 4\u00c2\u00b0C for 1 h. 50 \u00ce\u00bcl pre cleared embryo lysate was saved as Input and the left were incubated with affinity purified rabbit polyclonal anti Ybx1 antibody prepared by AbMax China and 30 \u00ce\u00bcl Protein A Dynabeads for 4 h at 4\u00c2\u00b0C or overnight. post washing with1 ml ice cold NT2 buffer 200 mM NaCl 50 mM HEPES pH 7.6 2 mM EDTA 0.05% NP 40 0.5 mM DTT 0.4U/\u00c2\u00b5l RNasin for eight times and once with 1ml ice cold 1\u00c3\u2014 PK buffer 100 mM Tris HCl pH 7.4 50 mM NaCl 10 mM EDTA 0.2% SDS the beads was treated in 200 ul PK buffer containing 20 ul proteinase K Roche 0311582001 for 1 h at 55\u00c2\u00b0C. The solution was collected and subjected to RNA extraction with Acid Phenol: ChCl3 pH4.34.7 and ethanol precipitation. The Input RNA was extracted by using TRIzol reagent. Both the Input and IP RNA were treated by TURBO\u00e2\u201e\u00a2 DNase Invitrogen AM2238. iCLIP seq: 1000 zebrafish embryos at 4 hpf were irradiated twice with 0.8 J/cm2 Stratalinker 2400 Stratagene lysed and subjected to mild fragmentation. Crosslinked RNA protein complexes were immunoprecipated using polyclonal Ybx1 antibody Abmax and protein A dynabeads. RNA was extracted and subjected to library construction using Smarter smRNA Seq kit Takara. All the libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 150 bp. RNA RIP BisSeq: Briefly 500 zebrafish embryos at shield stage 4 hpf were resuspended with 2 ml lysis buffer 150 mM KCl 10 mM HEPES pH 7.6 2 mM EDTA 0.5% NP 40 0.5 mM DTT 1:100 protease inhibitor cocktail 0.4 U/\u00ce\u00bcl RNasin by rotating at 4\u00c2\u00b0C for 30 min and centrifuged at 15 000 g for 15 min. The supernatant was collected and pre cleared with protein A Dynabeads by rotation at 4\u00c2\u00b0C for 1 h. 50 \u00ce\u00bcl pre cleared embryo lysate was saved as Input and the left were incubated with affinity purified rabbit polyclonal anti Ybx1 antibody prepared by AbMax China and 30 \u00ce\u00bcl Protein A Dynabeads for 4 h at 4\u00c2\u00b0C or overnight. post washing with1 ml ice cold NT2 buffer 200 mM NaCl 50 mM HEPES pH 7.6 2 mM EDTA 0.05% NP 40 0.5 mM DTT 0.4U/\u00c2\u00b5l RNasin for eight times and once with 1ml ice cold 1\u00c3\u2014 PK buffer 100 mM Tris HCl pH 7.4 50 mM NaCl 10 mM EDTA 0.2% SDS the beads was treated in 200 ul PK buffer containing 20 ul proteinase K Roche 0311582001 for 1 h at 55\u00c2\u00b0C. The solution was collected and subjected to RNA extraction with Acid Phenol: ChCl3 pH4.34.7 and ethanol precipitation. The purified mRNA was used for RNA BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 Dhfr mRNA serves as methylation conversion control was fragmented to 100 nt fragments for 1 min at 90\u00b0C in 10\u00d7 RNA Fragmentation Reagent Ambion then stopped by 10\u00d7 RNA stop solution Ambion and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 \u00b5l bisulfite solution pH 5.1 which is a 100:1 mixture of 40% sodium bisulfite Sigma and 600 \u00b5M hydroquinone Sigma and subjected to heat incubation at 75\u00b0C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns PALL Corporation with centrifugation. post washed with nuclease free water and centrifuged for five times the RNA was finally disolved in 75 ?l nuclease free water and then desulfonated by incubation with an equal volume of 1 M Tris HCl pH 9.0 at 75 \u00b0C for 1 h. post ethanol precipitation the RNA was resuspended in 11 \u00b5l of RNase free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase Invitrogen and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA Seq Kit KAPA according to the manufacturer?s instructions.Libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 125 bp.", "parent bioproject:PRJNA534030", null, null, "RIP BS 0hpf", "GSM3732426", null, "source name:Zebrafish embryo|strain:AB strain|age:0 hpf|tissue:whole embryo|rip antibody:polyclonal anti Ybx1 antibody", "RIP BS 0hpf", "library strategy: RNA RIP BisSeq Reads were aligned to the zv9 genome assembly using meRanTK v1.2.0 m5C sites were called by meRanCall v1.2.0 and annotated by applying BEDTools\u2019 intersectBed. MACS2 v2.1.1 were used for the peak calling and peaks were annotated by applying BEDTools\u2019 intersectBed. Genome build: zv9 Supplementary files format and content: Ybx1 binding m5C sites in one biological replicates.", "Zebrafish embryo", null, "Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol\u00ae Reagent Ambion. mRNA was extracted with using Dynabeads\u00ae mRNA Purification Kit Ambion and subjected to TURBO\u2122 DNase Invitrogen treatment at 37\u00b0C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. Thus purified mRNA was used for RNA BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 Dhfr mRNA serves as methylation conversion control was fragmented to 100 nt fragments for 1 min at 90\u00b0C in 10\u00d7 RNA Fragmentation Reagent Ambion then stopped by 10\u00d7 RNA stop solution Ambion and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 \u00b5l bisulfite solution pH 5.1 which is a 100:1 mixture of 40% sodium bisulfite Sigma and 600 \u00b5M hydroquinone Sigma and subjected to heat incubation at 75\u00b0C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns PALL Corporation with centrifugation. post washed with nuclease free water and centrifuged for five times the RNA was finally disolved in 75 \u03bcl nuclease free water and then desulfonated by incubation with an equal volume of 1 M Tris HCl pH 9.0 at 75 \u00b0C for 1 h. post ethanol precipitation the RNA was resuspended in 11 \u00b5l of RNase free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase Invitrogen and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA Seq Kit KAPA according to the manufacturer\u2019s instructions.Libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 125 bp.", null, "strain:AB strain|age:0 hpf|tissue:whole embryo|rip antibody:polyclonal anti Ybx1 antibody", "GSM3732426", "GSM3732426: RIP BS 0hpf; Danio rerio; OTHER", "GSM3732426", null, "1", "Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol\u00ae Reagent Ambion. mRNA was extracted with using Dynabeads\u00ae mRNA Purification Kit Ambion and subjected to TURBO\u2122 DNase Invitrogen treatment at 37\u00b0C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. Thus purified mRNA was used for RNA BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 Dhfr mRNA serves as methylation conversion control was fragmented to 100 nt fragments for 1 min at 90\u00b0C in 10\u00d7 RNA Fragmentation Reagent Ambion then stopped by 10\u00d7 RNA stop solution Ambion and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 \u00b5l bisulfite solution pH 5.1 which is a 100:1 mixture of 40% sodium bisulfite Sigma and 600 \u00b5M hydroquinone Sigma and subjected to heat incubation at 75\u00b0C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns PALL Corporation with centrifugation. post washed with nuclease free water and centrifuged for five times the RNA was finally disolved in 75 \u03bcl nuclease free water and then desulfonated by incubation with an equal volume of 1 M Tris HCl pH 9.0 at 75 \u00b0C for 1 h. post ethanol precipitation the RNA was resuspended in 11 \u00b5l of RNase free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase Invitrogen and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA Seq Kit KAPA according to the manufacturer's instructions.Libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 125 bp.", "GEO Accession:GSM3732426", "OTHER", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP162876", null, null, "RIP-BS_0hpf_R1.fastq.gz RIP-BS_0hpf_R2.fastq.gz", "fastq fastq", 41224496700.0, 137414989.0, "GSM3732426 r1", "0:150 1:150", "A:11515809359;C:9072060634;G:9856717145;T:10775122637;N:4786925", 150, 150, null, null, 11515809359, 9072060634, 9856717145, 10775122637, 4786925, "SRX5717519", "SRS4655939", "SRA786939", "GEO", "Beijing Institute of Genomics (BIG) of Chinese Academy of Sciences (CAS)", 2, 0.00575, 0.00421, 0.00122, 0.00083, 0.99738, 0.99801, 0.70652, 0.78053, 150, 150, "T", "T", "mates < 9% mapping rate", "illumina", "hiseq_era", "full_length", "random_priming", "smarter", "bulk", "clip", "iclip", null, "China", "2019-04-22", "Zygote", "Embryo", "Whole Organism", "All anatomical structures"], [49541, "SRR8937003", "SRX5717516", "SRS4655936", "SRP162876", "PRJNA493828", "Ybx1 binding sites and Ybx1 binding m5C sites in zebrafish embryo sample.", "GSE120646", "Other", "The maternal to zygotic transition MZT is a conserved and fundamental process during which the embryo undergoes dramatic reprogramming to convert maternal environment to embryonic driven programing. However how the maternally supplied transcripts are dynamically regulated during MZT remains largely unknown. Herein through genome wide profiling of RNA 5 methylcytosine m5C in zebrafish early embryos we show that m5C methylated maternal mRNAs display higher stability during MZT. We identify that the Y box binding protein 1 Ybx1 prefers to recognizing m5C modified mRNAs through p p interaction with a key residue Trp45 in its cold shock domain CSD which plays essential roles in maternal mRNA stability and early embryogenesis of zebrafish. Cooperated with an mRNA stabilizer Pabpc1a Ybx1 promotes the stability of its target mRNAs in an m5C dependent manner. Our study demonstrates a novel mechanism of RNA m5C methylation regulated maternal mRNA stability during zebrafish MZT highlighting the critical role of m5C mRNA methylation in early development. Overall design: Examination of Ybx1 binding sites and Ybx1 binding m5C sites in zebrafish embryo. RIP seq: Briefly 500 zebrafish embryos at shield stage 4 hpf were resuspended with 2 ml lysis buffer 150 mM KCl 10 mM HEPES pH 7.6 2 mM EDTA 0.5% NP 40 0.5 mM DTT 1:100 protease inhibitor cocktail 0.4 U/\u00ce\u00bcl RNasin by rotating at 4\u00c2\u00b0C for 30 min and centrifuged at 15 000 g for 15 min. The supernatant was collected and pre cleared with protein A Dynabeads by rotation at 4\u00c2\u00b0C for 1 h. 50 \u00ce\u00bcl pre cleared embryo lysate was saved as Input and the left were incubated with affinity purified rabbit polyclonal anti Ybx1 antibody prepared by AbMax China and 30 \u00ce\u00bcl Protein A Dynabeads for 4 h at 4\u00c2\u00b0C or overnight. post washing with1 ml ice cold NT2 buffer 200 mM NaCl 50 mM HEPES pH 7.6 2 mM EDTA 0.05% NP 40 0.5 mM DTT 0.4U/\u00c2\u00b5l RNasin for eight times and once with 1ml ice cold 1\u00c3\u2014 PK buffer 100 mM Tris HCl pH 7.4 50 mM NaCl 10 mM EDTA 0.2% SDS the beads was treated in 200 ul PK buffer containing 20 ul proteinase K Roche 0311582001 for 1 h at 55\u00c2\u00b0C. The solution was collected and subjected to RNA extraction with Acid Phenol: ChCl3 pH4.34.7 and ethanol precipitation. The Input RNA was extracted by using TRIzol reagent. Both the Input and IP RNA were treated by TURBO\u00e2\u201e\u00a2 DNase Invitrogen AM2238. iCLIP seq: 1000 zebrafish embryos at 4 hpf were irradiated twice with 0.8 J/cm2 Stratalinker 2400 Stratagene lysed and subjected to mild fragmentation. Crosslinked RNA protein complexes were immunoprecipated using polyclonal Ybx1 antibody Abmax and protein A dynabeads. RNA was extracted and subjected to library construction using Smarter smRNA Seq kit Takara. All the libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 150 bp. RNA RIP BisSeq: Briefly 500 zebrafish embryos at shield stage 4 hpf were resuspended with 2 ml lysis buffer 150 mM KCl 10 mM HEPES pH 7.6 2 mM EDTA 0.5% NP 40 0.5 mM DTT 1:100 protease inhibitor cocktail 0.4 U/\u00ce\u00bcl RNasin by rotating at 4\u00c2\u00b0C for 30 min and centrifuged at 15 000 g for 15 min. The supernatant was collected and pre cleared with protein A Dynabeads by rotation at 4\u00c2\u00b0C for 1 h. 50 \u00ce\u00bcl pre cleared embryo lysate was saved as Input and the left were incubated with affinity purified rabbit polyclonal anti Ybx1 antibody prepared by AbMax China and 30 \u00ce\u00bcl Protein A Dynabeads for 4 h at 4\u00c2\u00b0C or overnight. post washing with1 ml ice cold NT2 buffer 200 mM NaCl 50 mM HEPES pH 7.6 2 mM EDTA 0.05% NP 40 0.5 mM DTT 0.4U/\u00c2\u00b5l RNasin for eight times and once with 1ml ice cold 1\u00c3\u2014 PK buffer 100 mM Tris HCl pH 7.4 50 mM NaCl 10 mM EDTA 0.2% SDS the beads was treated in 200 ul PK buffer containing 20 ul proteinase K Roche 0311582001 for 1 h at 55\u00c2\u00b0C. The solution was collected and subjected to RNA extraction with Acid Phenol: ChCl3 pH4.34.7 and ethanol precipitation. The purified mRNA was used for RNA BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 Dhfr mRNA serves as methylation conversion control was fragmented to 100 nt fragments for 1 min at 90\u00b0C in 10\u00d7 RNA Fragmentation Reagent Ambion then stopped by 10\u00d7 RNA stop solution Ambion and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 \u00b5l bisulfite solution pH 5.1 which is a 100:1 mixture of 40% sodium bisulfite Sigma and 600 \u00b5M hydroquinone Sigma and subjected to heat incubation at 75\u00b0C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns PALL Corporation with centrifugation. post washed with nuclease free water and centrifuged for five times the RNA was finally disolved in 75 ?l nuclease free water and then desulfonated by incubation with an equal volume of 1 M Tris HCl pH 9.0 at 75 \u00b0C for 1 h. post ethanol precipitation the RNA was resuspended in 11 \u00b5l of RNase free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase Invitrogen and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA Seq Kit KAPA according to the manufacturer?s instructions.Libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 125 bp.", "parent bioproject:PRJNA534030", null, null, "RIP INPUT 0hpf rep2", "GSM3732423", null, "source name:RIP INPUT 0hpf|strain:AB strain|age:0 hpf|tissue:whole embryo", "RIP INPUT 0hpf rep2", "Reads were aligned to the zv9 genome assembly using TopHat v2.1.1. MACS2 v2.1.1 were used for the peak calling and peaks were annotated by applying BEDTools\u2019 intersectBed. Genome build: zv9 Supplementary files format and content: Ybx1 binding sites in two biological replicates.", "RIP INPUT 0hpf", null, "Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol\u00ae Reagent Ambion. mRNA was extracted with using Dynabeads\u00ae mRNA Purification Kit Ambion and subjected to TURBO\u2122 DNase Invitrogen treatment at 37\u00b0C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. RNA was extracted and subjected to library construction using Smarter smRNA Seq kit Takara. All the libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 150 bp.", null, "strain:AB strain|age:0 hpf|tissue:whole embryo", "GSM3732423", "GSM3732423: RIP INPUT 0hpf rep2; Danio rerio; RIP Seq", "GSM3732423", null, "1", "Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol\u00ae Reagent Ambion. mRNA was extracted with using Dynabeads\u00ae mRNA Purification Kit Ambion and subjected to TURBO\u2122 DNase Invitrogen treatment at 37\u00b0C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. RNA was extracted and subjected to library construction using Smarter smRNA Seq kit Takara. All the libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 150 bp.", "GEO Accession:GSM3732423", "RIP-Seq", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP162876", null, null, "RIP-INPUT_0hpf_rep2_R1.fastq.gz RIP-INPUT_0hpf_rep2_R2.fastq.gz", "fastq fastq", 17402218800.0, 58007396.0, "GSM3732423 r1", "0:150 1:150", "A:3416144403;C:5315174821;G:5345201218;T:3320906221;N:4792137", 150, 150, null, null, 3416144403, 5315174821, 5345201218, 3320906221, 4792137, "SRX5717516", "SRS4655936", "SRA786939", "GEO", "Beijing Institute of Genomics (BIG) of Chinese Academy of Sciences (CAS)", 2, 0.9819, 0.9814, 0.33605, 0.34016, 0.93458, 0.93963, 0.90172, 0.91272, 150, 150, "B", "B", "biological fallback assumption", "illumina", "hiseq_era", "full_length", "random_priming", "smarter", "bulk", "clip", "iclip", null, "China", "2019-04-22", "Zygote", "Embryo", "Whole Organism", "All anatomical structures"], [49542, "SRR8937002", "SRX5717515", "SRS4655935", "SRP162876", "PRJNA493828", "Ybx1 binding sites and Ybx1 binding m5C sites in zebrafish embryo sample.", "GSE120646", "Other", "The maternal to zygotic transition MZT is a conserved and fundamental process during which the embryo undergoes dramatic reprogramming to convert maternal environment to embryonic driven programing. However how the maternally supplied transcripts are dynamically regulated during MZT remains largely unknown. Herein through genome wide profiling of RNA 5 methylcytosine m5C in zebrafish early embryos we show that m5C methylated maternal mRNAs display higher stability during MZT. We identify that the Y box binding protein 1 Ybx1 prefers to recognizing m5C modified mRNAs through p p interaction with a key residue Trp45 in its cold shock domain CSD which plays essential roles in maternal mRNA stability and early embryogenesis of zebrafish. Cooperated with an mRNA stabilizer Pabpc1a Ybx1 promotes the stability of its target mRNAs in an m5C dependent manner. Our study demonstrates a novel mechanism of RNA m5C methylation regulated maternal mRNA stability during zebrafish MZT highlighting the critical role of m5C mRNA methylation in early development. Overall design: Examination of Ybx1 binding sites and Ybx1 binding m5C sites in zebrafish embryo. RIP seq: Briefly 500 zebrafish embryos at shield stage 4 hpf were resuspended with 2 ml lysis buffer 150 mM KCl 10 mM HEPES pH 7.6 2 mM EDTA 0.5% NP 40 0.5 mM DTT 1:100 protease inhibitor cocktail 0.4 U/\u00ce\u00bcl RNasin by rotating at 4\u00c2\u00b0C for 30 min and centrifuged at 15 000 g for 15 min. The supernatant was collected and pre cleared with protein A Dynabeads by rotation at 4\u00c2\u00b0C for 1 h. 50 \u00ce\u00bcl pre cleared embryo lysate was saved as Input and the left were incubated with affinity purified rabbit polyclonal anti Ybx1 antibody prepared by AbMax China and 30 \u00ce\u00bcl Protein A Dynabeads for 4 h at 4\u00c2\u00b0C or overnight. post washing with1 ml ice cold NT2 buffer 200 mM NaCl 50 mM HEPES pH 7.6 2 mM EDTA 0.05% NP 40 0.5 mM DTT 0.4U/\u00c2\u00b5l RNasin for eight times and once with 1ml ice cold 1\u00c3\u2014 PK buffer 100 mM Tris HCl pH 7.4 50 mM NaCl 10 mM EDTA 0.2% SDS the beads was treated in 200 ul PK buffer containing 20 ul proteinase K Roche 0311582001 for 1 h at 55\u00c2\u00b0C. The solution was collected and subjected to RNA extraction with Acid Phenol: ChCl3 pH4.34.7 and ethanol precipitation. The Input RNA was extracted by using TRIzol reagent. Both the Input and IP RNA were treated by TURBO\u00e2\u201e\u00a2 DNase Invitrogen AM2238. iCLIP seq: 1000 zebrafish embryos at 4 hpf were irradiated twice with 0.8 J/cm2 Stratalinker 2400 Stratagene lysed and subjected to mild fragmentation. Crosslinked RNA protein complexes were immunoprecipated using polyclonal Ybx1 antibody Abmax and protein A dynabeads. RNA was extracted and subjected to library construction using Smarter smRNA Seq kit Takara. All the libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 150 bp. RNA RIP BisSeq: Briefly 500 zebrafish embryos at shield stage 4 hpf were resuspended with 2 ml lysis buffer 150 mM KCl 10 mM HEPES pH 7.6 2 mM EDTA 0.5% NP 40 0.5 mM DTT 1:100 protease inhibitor cocktail 0.4 U/\u00ce\u00bcl RNasin by rotating at 4\u00c2\u00b0C for 30 min and centrifuged at 15 000 g for 15 min. The supernatant was collected and pre cleared with protein A Dynabeads by rotation at 4\u00c2\u00b0C for 1 h. 50 \u00ce\u00bcl pre cleared embryo lysate was saved as Input and the left were incubated with affinity purified rabbit polyclonal anti Ybx1 antibody prepared by AbMax China and 30 \u00ce\u00bcl Protein A Dynabeads for 4 h at 4\u00c2\u00b0C or overnight. post washing with1 ml ice cold NT2 buffer 200 mM NaCl 50 mM HEPES pH 7.6 2 mM EDTA 0.05% NP 40 0.5 mM DTT 0.4U/\u00c2\u00b5l RNasin for eight times and once with 1ml ice cold 1\u00c3\u2014 PK buffer 100 mM Tris HCl pH 7.4 50 mM NaCl 10 mM EDTA 0.2% SDS the beads was treated in 200 ul PK buffer containing 20 ul proteinase K Roche 0311582001 for 1 h at 55\u00c2\u00b0C. The solution was collected and subjected to RNA extraction with Acid Phenol: ChCl3 pH4.34.7 and ethanol precipitation. The purified mRNA was used for RNA BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 Dhfr mRNA serves as methylation conversion control was fragmented to 100 nt fragments for 1 min at 90\u00b0C in 10\u00d7 RNA Fragmentation Reagent Ambion then stopped by 10\u00d7 RNA stop solution Ambion and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 \u00b5l bisulfite solution pH 5.1 which is a 100:1 mixture of 40% sodium bisulfite Sigma and 600 \u00b5M hydroquinone Sigma and subjected to heat incubation at 75\u00b0C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns PALL Corporation with centrifugation. post washed with nuclease free water and centrifuged for five times the RNA was finally disolved in 75 ?l nuclease free water and then desulfonated by incubation with an equal volume of 1 M Tris HCl pH 9.0 at 75 \u00b0C for 1 h. post ethanol precipitation the RNA was resuspended in 11 \u00b5l of RNase free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase Invitrogen and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA Seq Kit KAPA according to the manufacturer?s instructions.Libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 125 bp.", "parent bioproject:PRJNA534030", null, null, "RIP INPUT 0hpf rep1", "GSM3732422", null, "source name:RIP INPUT 0hpf|strain:AB strain|age:0 hpf|tissue:whole embryo", "RIP INPUT 0hpf rep1", "Reads were aligned to the zv9 genome assembly using TopHat v2.1.1. MACS2 v2.1.1 were used for the peak calling and peaks were annotated by applying BEDTools\u2019 intersectBed. Genome build: zv9 Supplementary files format and content: Ybx1 binding sites in two biological replicates.", "RIP INPUT 0hpf", null, "Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol\u00ae Reagent Ambion. mRNA was extracted with using Dynabeads\u00ae mRNA Purification Kit Ambion and subjected to TURBO\u2122 DNase Invitrogen treatment at 37\u00b0C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. RNA was extracted and subjected to library construction using Smarter smRNA Seq kit Takara. All the libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 150 bp.", null, "strain:AB strain|age:0 hpf|tissue:whole embryo", "GSM3732422", "GSM3732422: RIP INPUT 0hpf rep1; Danio rerio; RIP Seq", "GSM3732422", null, "1", "Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol\u00ae Reagent Ambion. mRNA was extracted with using Dynabeads\u00ae mRNA Purification Kit Ambion and subjected to TURBO\u2122 DNase Invitrogen treatment at 37\u00b0C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. RNA was extracted and subjected to library construction using Smarter smRNA Seq kit Takara. All the libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 150 bp.", "GEO Accession:GSM3732422", "RIP-Seq", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP162876", null, null, "RIP-INPUT_0hpf_rep1_R1.fastq.gz RIP-INPUT_0hpf_rep1_R2.fastq.gz", "fastq fastq", 16615204800.0, 55384016.0, "GSM3732422 r1", "0:150 1:150", "A:2869477028;C:5392959051;G:5575878422;T:2776252538;N:637761", 150, 150, null, null, 2869477028, 5392959051, 5575878422, 2776252538, 637761, "SRX5717515", "SRS4655935", "SRA786939", "GEO", "Beijing Institute of Genomics (BIG) of Chinese Academy of Sciences (CAS)", 2, 0.97584, 0.96561, 0.04007, 0.03832, 0.9262, 0.933, 0.94642, 0.94376, 150, 150, "B", "B", "biological fallback assumption", "illumina", "hiseq_era", "full_length", "random_priming", "smarter", "bulk", "clip", "iclip", null, "China", "2019-04-22", "Zygote", "Embryo", "Whole Organism", "All anatomical structures"], [49545, "SRR8936999", "SRX5717512", "SRS4655932", "SRP162876", "PRJNA493828", "Ybx1 binding sites and Ybx1 binding m5C sites in zebrafish embryo sample.", "GSE120646", "Other", "The maternal to zygotic transition MZT is a conserved and fundamental process during which the embryo undergoes dramatic reprogramming to convert maternal environment to embryonic driven programing. However how the maternally supplied transcripts are dynamically regulated during MZT remains largely unknown. Herein through genome wide profiling of RNA 5 methylcytosine m5C in zebrafish early embryos we show that m5C methylated maternal mRNAs display higher stability during MZT. We identify that the Y box binding protein 1 Ybx1 prefers to recognizing m5C modified mRNAs through p p interaction with a key residue Trp45 in its cold shock domain CSD which plays essential roles in maternal mRNA stability and early embryogenesis of zebrafish. Cooperated with an mRNA stabilizer Pabpc1a Ybx1 promotes the stability of its target mRNAs in an m5C dependent manner. Our study demonstrates a novel mechanism of RNA m5C methylation regulated maternal mRNA stability during zebrafish MZT highlighting the critical role of m5C mRNA methylation in early development. Overall design: Examination of Ybx1 binding sites and Ybx1 binding m5C sites in zebrafish embryo. RIP seq: Briefly 500 zebrafish embryos at shield stage 4 hpf were resuspended with 2 ml lysis buffer 150 mM KCl 10 mM HEPES pH 7.6 2 mM EDTA 0.5% NP 40 0.5 mM DTT 1:100 protease inhibitor cocktail 0.4 U/\u00ce\u00bcl RNasin by rotating at 4\u00c2\u00b0C for 30 min and centrifuged at 15 000 g for 15 min. The supernatant was collected and pre cleared with protein A Dynabeads by rotation at 4\u00c2\u00b0C for 1 h. 50 \u00ce\u00bcl pre cleared embryo lysate was saved as Input and the left were incubated with affinity purified rabbit polyclonal anti Ybx1 antibody prepared by AbMax China and 30 \u00ce\u00bcl Protein A Dynabeads for 4 h at 4\u00c2\u00b0C or overnight. post washing with1 ml ice cold NT2 buffer 200 mM NaCl 50 mM HEPES pH 7.6 2 mM EDTA 0.05% NP 40 0.5 mM DTT 0.4U/\u00c2\u00b5l RNasin for eight times and once with 1ml ice cold 1\u00c3\u2014 PK buffer 100 mM Tris HCl pH 7.4 50 mM NaCl 10 mM EDTA 0.2% SDS the beads was treated in 200 ul PK buffer containing 20 ul proteinase K Roche 0311582001 for 1 h at 55\u00c2\u00b0C. The solution was collected and subjected to RNA extraction with Acid Phenol: ChCl3 pH4.34.7 and ethanol precipitation. The Input RNA was extracted by using TRIzol reagent. Both the Input and IP RNA were treated by TURBO\u00e2\u201e\u00a2 DNase Invitrogen AM2238. iCLIP seq: 1000 zebrafish embryos at 4 hpf were irradiated twice with 0.8 J/cm2 Stratalinker 2400 Stratagene lysed and subjected to mild fragmentation. Crosslinked RNA protein complexes were immunoprecipated using polyclonal Ybx1 antibody Abmax and protein A dynabeads. RNA was extracted and subjected to library construction using Smarter smRNA Seq kit Takara. All the libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 150 bp. RNA RIP BisSeq: Briefly 500 zebrafish embryos at shield stage 4 hpf were resuspended with 2 ml lysis buffer 150 mM KCl 10 mM HEPES pH 7.6 2 mM EDTA 0.5% NP 40 0.5 mM DTT 1:100 protease inhibitor cocktail 0.4 U/\u00ce\u00bcl RNasin by rotating at 4\u00c2\u00b0C for 30 min and centrifuged at 15 000 g for 15 min. The supernatant was collected and pre cleared with protein A Dynabeads by rotation at 4\u00c2\u00b0C for 1 h. 50 \u00ce\u00bcl pre cleared embryo lysate was saved as Input and the left were incubated with affinity purified rabbit polyclonal anti Ybx1 antibody prepared by AbMax China and 30 \u00ce\u00bcl Protein A Dynabeads for 4 h at 4\u00c2\u00b0C or overnight. post washing with1 ml ice cold NT2 buffer 200 mM NaCl 50 mM HEPES pH 7.6 2 mM EDTA 0.05% NP 40 0.5 mM DTT 0.4U/\u00c2\u00b5l RNasin for eight times and once with 1ml ice cold 1\u00c3\u2014 PK buffer 100 mM Tris HCl pH 7.4 50 mM NaCl 10 mM EDTA 0.2% SDS the beads was treated in 200 ul PK buffer containing 20 ul proteinase K Roche 0311582001 for 1 h at 55\u00c2\u00b0C. The solution was collected and subjected to RNA extraction with Acid Phenol: ChCl3 pH4.34.7 and ethanol precipitation. The purified mRNA was used for RNA BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 Dhfr mRNA serves as methylation conversion control was fragmented to 100 nt fragments for 1 min at 90\u00b0C in 10\u00d7 RNA Fragmentation Reagent Ambion then stopped by 10\u00d7 RNA stop solution Ambion and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 \u00b5l bisulfite solution pH 5.1 which is a 100:1 mixture of 40% sodium bisulfite Sigma and 600 \u00b5M hydroquinone Sigma and subjected to heat incubation at 75\u00b0C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns PALL Corporation with centrifugation. post washed with nuclease free water and centrifuged for five times the RNA was finally disolved in 75 ?l nuclease free water and then desulfonated by incubation with an equal volume of 1 M Tris HCl pH 9.0 at 75 \u00b0C for 1 h. post ethanol precipitation the RNA was resuspended in 11 \u00b5l of RNase free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase Invitrogen and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA Seq Kit KAPA according to the manufacturer?s instructions.Libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 125 bp.", "parent bioproject:PRJNA534030", null, null, "RIP 0hpf rep2", "GSM3732419", null, "source name:RIP 0hpf|strain:AB strain|age:0 hpf|tissue:whole embryo|rip antibody:rabbit polyclonal anti Ybx1 antibody", "RIP 0hpf rep2", "Reads were aligned to the zv9 genome assembly using TopHat v2.1.1. MACS2 v2.1.1 were used for the peak calling and peaks were annotated by applying BEDTools\u2019 intersectBed. Genome build: zv9 Supplementary files format and content: Ybx1 binding sites in two biological replicates.", "RIP 0hpf", null, "Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol\u00ae Reagent Ambion. mRNA was extracted with using Dynabeads\u00ae mRNA Purification Kit Ambion and subjected to TURBO\u2122 DNase Invitrogen treatment at 37\u00b0C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. RNA was extracted and subjected to library construction using Smarter smRNA Seq kit Takara. All the libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 150 bp.", null, "strain:AB strain|age:0 hpf|tissue:whole embryo|rip antibody:rabbit polyclonal anti Ybx1 antibody", "GSM3732419", "GSM3732419: RIP 0hpf rep2; Danio rerio; RIP Seq", "GSM3732419", null, "1", "Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol\u00ae Reagent Ambion. mRNA was extracted with using Dynabeads\u00ae mRNA Purification Kit Ambion and subjected to TURBO\u2122 DNase Invitrogen treatment at 37\u00b0C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. RNA was extracted and subjected to library construction using Smarter smRNA Seq kit Takara. All the libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 150 bp.", "GEO Accession:GSM3732419", "RIP-Seq", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP162876", null, null, "RIP_0hpf_rep2_R1.fastq.gz RIP_0hpf_rep2_R2.fastq.gz", "fastq fastq", 14319153300.0, 47730511.0, "GSM3732419 r1", "0:150 1:150", "A:3633266473;C:3581763715;G:3602109377;T:3500497801;N:1515934", 150, 150, null, null, 3633266473, 3581763715, 3602109377, 3500497801, 1515934, "SRX5717512", "SRS4655932", "SRA786939", "GEO", "Beijing Institute of Genomics (BIG) of Chinese Academy of Sciences (CAS)", 2, 0.96309, 0.96299, 0.09978, 0.09922, 0.80012, 0.80635, 0.7641, 0.76535, 150, 150, "B", "B", "biological fallback assumption", "illumina", "hiseq_era", "full_length", "random_priming", "smarter", "bulk", "clip", "iclip", null, "China", "2019-04-22", "Zygote", "Embryo", "Whole Organism", "All anatomical structures"], [49546, "SRR8936998", "SRX5717511", "SRS4655931", "SRP162876", "PRJNA493828", "Ybx1 binding sites and Ybx1 binding m5C sites in zebrafish embryo sample.", "GSE120646", "Other", "The maternal to zygotic transition MZT is a conserved and fundamental process during which the embryo undergoes dramatic reprogramming to convert maternal environment to embryonic driven programing. However how the maternally supplied transcripts are dynamically regulated during MZT remains largely unknown. Herein through genome wide profiling of RNA 5 methylcytosine m5C in zebrafish early embryos we show that m5C methylated maternal mRNAs display higher stability during MZT. We identify that the Y box binding protein 1 Ybx1 prefers to recognizing m5C modified mRNAs through p p interaction with a key residue Trp45 in its cold shock domain CSD which plays essential roles in maternal mRNA stability and early embryogenesis of zebrafish. Cooperated with an mRNA stabilizer Pabpc1a Ybx1 promotes the stability of its target mRNAs in an m5C dependent manner. Our study demonstrates a novel mechanism of RNA m5C methylation regulated maternal mRNA stability during zebrafish MZT highlighting the critical role of m5C mRNA methylation in early development. Overall design: Examination of Ybx1 binding sites and Ybx1 binding m5C sites in zebrafish embryo. RIP seq: Briefly 500 zebrafish embryos at shield stage 4 hpf were resuspended with 2 ml lysis buffer 150 mM KCl 10 mM HEPES pH 7.6 2 mM EDTA 0.5% NP 40 0.5 mM DTT 1:100 protease inhibitor cocktail 0.4 U/\u00ce\u00bcl RNasin by rotating at 4\u00c2\u00b0C for 30 min and centrifuged at 15 000 g for 15 min. The supernatant was collected and pre cleared with protein A Dynabeads by rotation at 4\u00c2\u00b0C for 1 h. 50 \u00ce\u00bcl pre cleared embryo lysate was saved as Input and the left were incubated with affinity purified rabbit polyclonal anti Ybx1 antibody prepared by AbMax China and 30 \u00ce\u00bcl Protein A Dynabeads for 4 h at 4\u00c2\u00b0C or overnight. post washing with1 ml ice cold NT2 buffer 200 mM NaCl 50 mM HEPES pH 7.6 2 mM EDTA 0.05% NP 40 0.5 mM DTT 0.4U/\u00c2\u00b5l RNasin for eight times and once with 1ml ice cold 1\u00c3\u2014 PK buffer 100 mM Tris HCl pH 7.4 50 mM NaCl 10 mM EDTA 0.2% SDS the beads was treated in 200 ul PK buffer containing 20 ul proteinase K Roche 0311582001 for 1 h at 55\u00c2\u00b0C. The solution was collected and subjected to RNA extraction with Acid Phenol: ChCl3 pH4.34.7 and ethanol precipitation. The Input RNA was extracted by using TRIzol reagent. Both the Input and IP RNA were treated by TURBO\u00e2\u201e\u00a2 DNase Invitrogen AM2238. iCLIP seq: 1000 zebrafish embryos at 4 hpf were irradiated twice with 0.8 J/cm2 Stratalinker 2400 Stratagene lysed and subjected to mild fragmentation. Crosslinked RNA protein complexes were immunoprecipated using polyclonal Ybx1 antibody Abmax and protein A dynabeads. RNA was extracted and subjected to library construction using Smarter smRNA Seq kit Takara. All the libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 150 bp. RNA RIP BisSeq: Briefly 500 zebrafish embryos at shield stage 4 hpf were resuspended with 2 ml lysis buffer 150 mM KCl 10 mM HEPES pH 7.6 2 mM EDTA 0.5% NP 40 0.5 mM DTT 1:100 protease inhibitor cocktail 0.4 U/\u00ce\u00bcl RNasin by rotating at 4\u00c2\u00b0C for 30 min and centrifuged at 15 000 g for 15 min. The supernatant was collected and pre cleared with protein A Dynabeads by rotation at 4\u00c2\u00b0C for 1 h. 50 \u00ce\u00bcl pre cleared embryo lysate was saved as Input and the left were incubated with affinity purified rabbit polyclonal anti Ybx1 antibody prepared by AbMax China and 30 \u00ce\u00bcl Protein A Dynabeads for 4 h at 4\u00c2\u00b0C or overnight. post washing with1 ml ice cold NT2 buffer 200 mM NaCl 50 mM HEPES pH 7.6 2 mM EDTA 0.05% NP 40 0.5 mM DTT 0.4U/\u00c2\u00b5l RNasin for eight times and once with 1ml ice cold 1\u00c3\u2014 PK buffer 100 mM Tris HCl pH 7.4 50 mM NaCl 10 mM EDTA 0.2% SDS the beads was treated in 200 ul PK buffer containing 20 ul proteinase K Roche 0311582001 for 1 h at 55\u00c2\u00b0C. The solution was collected and subjected to RNA extraction with Acid Phenol: ChCl3 pH4.34.7 and ethanol precipitation. The purified mRNA was used for RNA BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 Dhfr mRNA serves as methylation conversion control was fragmented to 100 nt fragments for 1 min at 90\u00b0C in 10\u00d7 RNA Fragmentation Reagent Ambion then stopped by 10\u00d7 RNA stop solution Ambion and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 \u00b5l bisulfite solution pH 5.1 which is a 100:1 mixture of 40% sodium bisulfite Sigma and 600 \u00b5M hydroquinone Sigma and subjected to heat incubation at 75\u00b0C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns PALL Corporation with centrifugation. post washed with nuclease free water and centrifuged for five times the RNA was finally disolved in 75 ?l nuclease free water and then desulfonated by incubation with an equal volume of 1 M Tris HCl pH 9.0 at 75 \u00b0C for 1 h. post ethanol precipitation the RNA was resuspended in 11 \u00b5l of RNase free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase Invitrogen and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA Seq Kit KAPA according to the manufacturer?s instructions.Libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 125 bp.", "parent bioproject:PRJNA534030", null, null, "RIP 0hpf rep1", "GSM3732418", null, "source name:RIP 0hpf|strain:AB strain|age:0 hpf|tissue:whole embryo|rip antibody:rabbit polyclonal anti Ybx1 antibody", "RIP 0hpf rep1", "Reads were aligned to the zv9 genome assembly using TopHat v2.1.1. MACS2 v2.1.1 were used for the peak calling and peaks were annotated by applying BEDTools\u2019 intersectBed. Genome build: zv9 Supplementary files format and content: Ybx1 binding sites in two biological replicates.", "RIP 0hpf", null, "Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol\u00ae Reagent Ambion. mRNA was extracted with using Dynabeads\u00ae mRNA Purification Kit Ambion and subjected to TURBO\u2122 DNase Invitrogen treatment at 37\u00b0C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. RNA was extracted and subjected to library construction using Smarter smRNA Seq kit Takara. All the libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 150 bp.", null, "strain:AB strain|age:0 hpf|tissue:whole embryo|rip antibody:rabbit polyclonal anti Ybx1 antibody", "GSM3732418", "GSM3732418: RIP 0hpf rep1; Danio rerio; RIP Seq", "GSM3732418", null, "1", "Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol\u00ae Reagent Ambion. mRNA was extracted with using Dynabeads\u00ae mRNA Purification Kit Ambion and subjected to TURBO\u2122 DNase Invitrogen treatment at 37\u00b0C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. RNA was extracted and subjected to library construction using Smarter smRNA Seq kit Takara. All the libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 150 bp.", "GEO Accession:GSM3732418", "RIP-Seq", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP162876", null, null, "RIP_0hpf_rep1_R1.fastq.gz RIP_0hpf_rep1_R2.fastq.gz", "fastq fastq", 7807878000.0, 26026260.0, "GSM3732418 r1", "0:150 1:150", "A:1698013251;C:2200605244;G:2305251364;T:1602824248;N:1183893", 150, 150, null, null, 1698013251, 2200605244, 2305251364, 1602824248, 1183893, "SRX5717511", "SRS4655931", "SRA786939", "GEO", "Beijing Institute of Genomics (BIG) of Chinese Academy of Sciences (CAS)", 2, 0.89154, 0.89105, 0.26373, 0.26886, 0.82477, 0.827, 0.79125, 0.77711, 150, 150, "B", "B", "biological fallback assumption", "illumina", "hiseq_era", "full_length", "random_priming", "smarter", "bulk", "clip", "iclip", null, "China", "2019-04-22", "Zygote", "Embryo", "Whole Organism", "All anatomical structures"], [49563, "SRR7942622", "SRX4776887", "SRS3857428", "SRP162875", "PRJNA493829", "m5C profiles in zebrafish embryo sample.", "GSE120645", "Other", "Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol\u00ae Reagent Ambion. mRNA was extracted with using Dynabeads\u00ae mRNA Purification Kit Ambion and subjected to TURBO\u2122 DNase Invitrogen treatment at 37\u00b0C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 Dhfr mRNA serves as methylation conversion control was fragmented to 100 nt fragments for 1 min at 90\u00b0C in 10\u00d7 RNA Fragmentation Reagent Ambion then stopped by 10\u00d7 RNA stop solution Ambion and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 \u00b5l bisulfite solution pH 5.1 which is a 100:1 mixture of 40% sodium bisulfite Sigma and 600 \u00b5M hydroquinone Sigma and subjected to heat incubation at 75\u00b0C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns PALL Corporation with centrifugation. post washed with nuclease free water and centrifuged for five times the RNA was finally disolved in 75 \u00b5l nuclease free water and then desulfonated by incubation with an equal volume of 1 M Tris HCl pH 9.0 at 75 \u00b0C for 1 h. post ethanol precipitation the RNA was resuspended in 11 \u00b5l of RNase free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase Invitrogen and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA Seq Kit KAPA according to the manufacturer's instructions.Libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 125 bp. Overall design: Examination of m5C levels in zebrafish embryo.", null, null, null, "WT 0hpf m5C rep2", "GSM3406888", null, "source name:Zebrafish embryo|strain:AB strain|cell type:Zebrafish embryo|age:0 hpf|tissue:whole embryo", "WT 0hpf m5C rep2", "Reads were aligned to the zv9 and hg19 genome assembly using meRanTK v1.2.0 m5C sites were called by meRanCall v1.2.0 and annotated by applying BEDTools\u2019 intersectBed. Genome build: zv9 and hg19 Supplementary files format and content: m5C sites in two biological replicates.", "Zebrafish embryo", null, "Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol\u00ae Reagent Ambion. mRNA was extracted with using Dynabeads\u00ae mRNA Purification Kit Ambion and subjected to TURBO\u2122 DNase Invitrogen treatment at 37\u00b0C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. Thus purified mRNA was used for RNA BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 Dhfr mRNA serves as methylation conversion control was fragmented to 100 nt fragments for 1 min at 90\u00b0C in 10\u00d7 RNA Fragmentation Reagent Ambion then stopped by 10\u00d7 RNA stop solution Ambion and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 \u00b5l bisulfite solution pH 5.1 which is a 100:1 mixture of 40% sodium bisulfite Sigma and 600 \u00b5M hydroquinone Sigma and subjected to heat incubation at 75\u00b0C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns PALL Corporation with centrifugation. post washed with nuclease free water and centrifuged for five times the RNA was finally disolved in 75 \u03bcl nuclease free water and then desulfonated by incubation with an equal volume of 1 M Tris HCl pH 9.0 at 75 \u00b0C for 1 h. post ethanol precipitation the RNA was resuspended in 11 \u00b5l of RNase free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase Invitrogen and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA Seq Kit KAPA according to the manufacturer\u2019s instructions.Libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 125 bp. RNA BisSeq", null, "strain:AB strain|cell type:Zebrafish embryo|age:0 hpf|tissue:whole embryo", "GSM3406888", "GSM3406888: WT 0hpf m5C rep2; Danio rerio; OTHER", "GSM3406888", null, "1", "Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol\u00ae Reagent Ambion. mRNA was extracted with using Dynabeads\u00ae mRNA Purification Kit Ambion and subjected to TURBO\u2122 DNase Invitrogen treatment at 37\u00b0C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. Thus purified mRNA was used for RNA BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 Dhfr mRNA serves as methylation conversion control was fragmented to 100 nt fragments for 1 min at 90\u00b0C in 10\u00d7 RNA Fragmentation Reagent Ambion then stopped by 10\u00d7 RNA stop solution Ambion and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 \u00b5l bisulfite solution pH 5.1 which is a 100:1 mixture of 40% sodium bisulfite Sigma and 600 \u00b5M hydroquinone Sigma and subjected to heat incubation at 75\u00b0C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns PALL Corporation with centrifugation. post washed with nuclease free water and centrifuged for five times the RNA was finally disolved in 75 \u03bcl nuclease free water and then desulfonated by incubation with an equal volume of 1 M Tris HCl pH 9.0 at 75 \u00b0C for 1 h. post ethanol precipitation the RNA was resuspended in 11 \u00b5l of RNase free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase Invitrogen and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA Seq Kit KAPA according to the manufacturer's instructions.Libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 125 bp. RNA BisSeq", "GEO Accession:GSM3406888", "OTHER", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP162875", null, null, "WT_0hpf_rep2_m5C_R1.fastq.gz WT_0hpf_rep2_m5C_R2.fastq.gz", "fastq fastq", 21201784250.0, 84807137.0, "GSM3406888 r1", "0:125 1:125", "A:7176675495;C:3469343240;G:3951404122;T:6579575790;N:24785603", 125, 125, null, null, 7176675495, 3469343240, 3951404122, 6579575790, 24785603, "SRX4776887", "SRS3857428", "SRA786937", "GEO", "Beijing Institute of Genomics (BIG) of Chinese Academy of Sciences (CAS)", 2, 0.00074, 0.00077, 0.00015, 0.00015, 0.99835, 0.99829, 0.62037, 0.64035, 125, 125, "T", "T", "mates < 9% mapping rate", "illumina", "hiseq_era", "unknown", "random_priming", "unknown", "bulk", "unknown", "unknown", null, "China", "2018-09-28", "Zygote", "Embryo", "Whole Organism", "All anatomical structures"], [49564, "SRR7942621", "SRX4776886", "SRS3857426", "SRP162875", "PRJNA493829", "m5C profiles in zebrafish embryo sample.", "GSE120645", "Other", "Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol\u00ae Reagent Ambion. mRNA was extracted with using Dynabeads\u00ae mRNA Purification Kit Ambion and subjected to TURBO\u2122 DNase Invitrogen treatment at 37\u00b0C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 Dhfr mRNA serves as methylation conversion control was fragmented to 100 nt fragments for 1 min at 90\u00b0C in 10\u00d7 RNA Fragmentation Reagent Ambion then stopped by 10\u00d7 RNA stop solution Ambion and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 \u00b5l bisulfite solution pH 5.1 which is a 100:1 mixture of 40% sodium bisulfite Sigma and 600 \u00b5M hydroquinone Sigma and subjected to heat incubation at 75\u00b0C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns PALL Corporation with centrifugation. post washed with nuclease free water and centrifuged for five times the RNA was finally disolved in 75 \u00b5l nuclease free water and then desulfonated by incubation with an equal volume of 1 M Tris HCl pH 9.0 at 75 \u00b0C for 1 h. post ethanol precipitation the RNA was resuspended in 11 \u00b5l of RNase free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase Invitrogen and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA Seq Kit KAPA according to the manufacturer's instructions.Libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 125 bp. Overall design: Examination of m5C levels in zebrafish embryo.", null, null, null, "WT 0hpf m5C rep1", "GSM3406887", null, "source name:Zebrafish embryo|strain:AB strain|cell type:Zebrafish embryo|age:0 hpf|tissue:whole embryo", "WT 0hpf m5C rep1", "Reads were aligned to the zv9 and hg19 genome assembly using meRanTK v1.2.0 m5C sites were called by meRanCall v1.2.0 and annotated by applying BEDTools\u2019 intersectBed. Genome build: zv9 and hg19 Supplementary files format and content: m5C sites in two biological replicates.", "Zebrafish embryo", null, "Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol\u00ae Reagent Ambion. mRNA was extracted with using Dynabeads\u00ae mRNA Purification Kit Ambion and subjected to TURBO\u2122 DNase Invitrogen treatment at 37\u00b0C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. Thus purified mRNA was used for RNA BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 Dhfr mRNA serves as methylation conversion control was fragmented to 100 nt fragments for 1 min at 90\u00b0C in 10\u00d7 RNA Fragmentation Reagent Ambion then stopped by 10\u00d7 RNA stop solution Ambion and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 \u00b5l bisulfite solution pH 5.1 which is a 100:1 mixture of 40% sodium bisulfite Sigma and 600 \u00b5M hydroquinone Sigma and subjected to heat incubation at 75\u00b0C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns PALL Corporation with centrifugation. post washed with nuclease free water and centrifuged for five times the RNA was finally disolved in 75 \u03bcl nuclease free water and then desulfonated by incubation with an equal volume of 1 M Tris HCl pH 9.0 at 75 \u00b0C for 1 h. post ethanol precipitation the RNA was resuspended in 11 \u00b5l of RNase free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase Invitrogen and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA Seq Kit KAPA according to the manufacturer\u2019s instructions.Libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 125 bp. RNA BisSeq", null, "strain:AB strain|cell type:Zebrafish embryo|age:0 hpf|tissue:whole embryo", "GSM3406887", "GSM3406887: WT 0hpf m5C rep1; Danio rerio; OTHER", "GSM3406887", null, "1", "Total RNA was isolated from zebrafish embryos harvested at different stages using TRIzol\u00ae Reagent Ambion. mRNA was extracted with using Dynabeads\u00ae mRNA Purification Kit Ambion and subjected to TURBO\u2122 DNase Invitrogen treatment at 37\u00b0C for 30 min and ethanol precipitation. Thus purified mRNA was used for RNA BisSeq library construction. Thus purified mRNA was used for RNA BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 Dhfr mRNA serves as methylation conversion control was fragmented to 100 nt fragments for 1 min at 90\u00b0C in 10\u00d7 RNA Fragmentation Reagent Ambion then stopped by 10\u00d7 RNA stop solution Ambion and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 \u00b5l bisulfite solution pH 5.1 which is a 100:1 mixture of 40% sodium bisulfite Sigma and 600 \u00b5M hydroquinone Sigma and subjected to heat incubation at 75\u00b0C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns PALL Corporation with centrifugation. post washed with nuclease free water and centrifuged for five times the RNA was finally disolved in 75 \u03bcl nuclease free water and then desulfonated by incubation with an equal volume of 1 M Tris HCl pH 9.0 at 75 \u00b0C for 1 h. post ethanol precipitation the RNA was resuspended in 11 \u00b5l of RNase free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase Invitrogen and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA Seq Kit KAPA according to the manufacturer's instructions.Libraries were sequenced using HiSeq2500 Illumina in paired read mode creating reads with a length of 125 bp. RNA BisSeq", "GEO Accession:GSM3406887", "OTHER", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP162875", null, null, "WT_0hpf_rep1_m5C_R1.fastq.gz WT_0hpf_rep1_m5C_R2.fastq.gz", "fastq fastq", 25405235500.0, 101620942.0, "GSM3406887 r1", "0:125 1:125", "A:8768618269;C:3912734696;G:4276708166;T:8422886588;N:24287781", 125, 125, null, null, 8768618269, 3912734696, 4276708166, 8422886588, 24287781, "SRX4776886", "SRS3857426", "SRA786937", "GEO", "Beijing Institute of Genomics (BIG) of Chinese Academy of Sciences (CAS)", 2, 0.0011, 0.00103, 0.00022, 0.00018, 0.9977, 0.99786, 0.60493, 0.60256, 125, 125, "T", "T", "mates < 9% mapping rate", "illumina", "hiseq_era", "unknown", "random_priming", "unknown", "bulk", "unknown", "unknown", null, "China", "2018-09-28", "Zygote", "Embryo", "Whole Organism", "All anatomical structures"], [49603, "SRR7947895", "SRX4781855", "SRS3862047", "SRP163087", "PRJNA494251", "Dynamic landscapes and regulome of mRNA structure during zebrafish early embryogenesis", "GSE120724", "Other", "We resolved the RNA secondary structure during zebrafish early embryogenesis based on in vivo click selective 2' hydroxyl acylation and profiling experiment icSHAPE. We analyzed the RNA structure dynamics among different development stages. Also we studied which factors regulate maternal gene decay by RNA structure switch. Overall design: icSHAPE of 12 samples from zebrafish embryos at six different early development stages and 4 samples from Elavl1a deficient zebrafish embryos at sphere and shield stages including two treated with DMSO as control and two treated with NAIN3 for each stage or condition. RNA seq of 6 samples from zebrafish embryos at 2 4 6 h.p.f. and 4 samples from Elavl1a deficient zebrafish embryos at 4 and 6 h.p.f. including two biological replicates. iCLIP seq of 4 samples from flag elavl1a mRNA injected zebrafish embryos at xxx and 6 h.p.f. including two biological replicates.", null, "pubmed:32423473", null, "1 cell NAIN3 icSHAPE", "GSM3409379", null, "source name:1 cell NAIN3|strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:0.4hpf", "1 cell NAIN3 icSHAPE", "Library strategy: icSHAPE icSHAPE computational pipeline reference: PubMedID: 26766114 DOI: 10.1038/nprot.2016.011 trimmed three prime adaptor from reads by cutadapt trimmed low quality bases by trimmomatic collapsed duplicated reads as icSHAPE pipeline trimmed the leading 13nt UMI mapping reads to zebrafish reference transcriptomez10 by bowtie2 using the same parameters in icSHAPE pipeline calculate rpkm as icshape pipeline calculate reverse transcription stop sitesRT stop as icshape pipeline RT stop was normalized by using sliding window then icSHAPE score was calculated as icshape pipeline Genome build: z10 Supplementary files format and content: tab delimited text files for icshape files. First column is the Refseq ID second is transcript length third is RPKM then every column is the nucleotide resolution icSHAPE score. NULL means no confident score. Tab delimited text files for read counts and rpkm files. First column is the Refseq ID second is transcript length third is the unique mapped reads count fourth is the mulitple mapped reads count fifth is the RPKM.", "1 cell NAIN3", "For in vivo treatment cells was incubated with NAI N3 or DMSO at 28.5 C for 5min.", "The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol\u00ae Reagent invitrogen.To enrich polyA RNA the total RNA was subjected to polyA selection. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37\u00b0C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/\u00b5l RiboLock 0.1 U/\u00b5l FastAP Life Technology 2 U/\u00b5l T4 PNKNEB before 3\u2019 adapter ligation with addition of 10 \u00b5l ligation mix 5mM DTT 0.5\u00b5M 5\u2019 adenylated and 3\u2019 blocked linker 3\u2019 bio for unmodified 3\u2019 ddc for modified 0.66 U/\u00b5l T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25\u00b0C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1\u00b5l FastAP 0.2\u00b5l SSB Promega 0.8\u00b5l Ribolock and 1\u00b5l 5\u2019 Deadenylase NEB in 1X NEB buffer 2 NEB at 30\u00b0C for 90 min. And then 1\u00b5l RecJf NEB was added with another incubation at 37\u00b0C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq\uff0c Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer\u2019s protocol.", "Zebrafish wild type strain AB was raised in system water at 28.5\u00b0C under standard conditions.", "strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:0.4hpf", "GSM3409379", "GSM3409379: 1 cell NAIN3; Danio rerio; OTHER", "GSM3409379", null, "1", "The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol\u00ae Reagent invitrogen. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37\u00b0C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/\u00b5l RiboLock 0.1 U/\u00b5l FastAP Life Technology 2 U/\u00b5l T4 PNKNEB before three prime adapter ligation with addition of 10 \u00b5l ligation mix 5mM DTT 0.5\u00b5M five prime adenylated and three prime blocked linker three prime bio for unmodified three prime ddc for modified 0.66 U/\u00b5l T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25\u00b0C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1\u00b5l FastAP 0.2\u00b5l SSB Promega 0.8\u00b5l Ribolock and 1\u00b5l five prime Deadenylase NEB in 1X NEB buffer 2 NEB at 30\u00b0C for 90 min. And then 1\u00b5l RecJf NEB was added with another incubation at 37\u00b0C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq\uff0c Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer's protocol.", "GEO Accession:GSM3409379", "OTHER", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP163087", null, null, "1-cell_NAIN3_rep1.fastq.gz", "fastq", 99090115350.0, 660600769.0, "GSM3409379 r1", "0:150 1:0", "A:27532157732;C:24798945523;G:22046350479;T:24700708479;N:11953137", 150, 0, null, null, 27532157732, 24798945523, 22046350479, 24700708479, 11953137, "SRX4781855", "SRS3862047", "SRA787572", "GEO", "Life Science, Tsinghua University", 1, 0.02566, null, 0.0042, null, 0.95467, null, 0.70522, null, 150, null, "B", null, "usable mapping rate", "illumina", "hiseq_era", "unknown", "poly_a", "unknown", "bulk", "clip", "iclip", null, "China", "2018-10-01", "Zygote", "Embryo", "Embryo Imprecise", "All anatomical structures"], [49604, "SRR7947896", "SRX4781855", "SRS3862047", "SRP163087", "PRJNA494251", "Dynamic landscapes and regulome of mRNA structure during zebrafish early embryogenesis", "GSE120724", "Other", "We resolved the RNA secondary structure during zebrafish early embryogenesis based on in vivo click selective 2' hydroxyl acylation and profiling experiment icSHAPE. We analyzed the RNA structure dynamics among different development stages. Also we studied which factors regulate maternal gene decay by RNA structure switch. Overall design: icSHAPE of 12 samples from zebrafish embryos at six different early development stages and 4 samples from Elavl1a deficient zebrafish embryos at sphere and shield stages including two treated with DMSO as control and two treated with NAIN3 for each stage or condition. RNA seq of 6 samples from zebrafish embryos at 2 4 6 h.p.f. and 4 samples from Elavl1a deficient zebrafish embryos at 4 and 6 h.p.f. including two biological replicates. iCLIP seq of 4 samples from flag elavl1a mRNA injected zebrafish embryos at xxx and 6 h.p.f. including two biological replicates.", null, "pubmed:32423473", null, "1 cell NAIN3 icSHAPE", "GSM3409379", null, "source name:1 cell NAIN3|strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:0.4hpf", "1 cell NAIN3 icSHAPE", "Library strategy: icSHAPE icSHAPE computational pipeline reference: PubMedID: 26766114 DOI: 10.1038/nprot.2016.011 trimmed three prime adaptor from reads by cutadapt trimmed low quality bases by trimmomatic collapsed duplicated reads as icSHAPE pipeline trimmed the leading 13nt UMI mapping reads to zebrafish reference transcriptomez10 by bowtie2 using the same parameters in icSHAPE pipeline calculate rpkm as icshape pipeline calculate reverse transcription stop sitesRT stop as icshape pipeline RT stop was normalized by using sliding window then icSHAPE score was calculated as icshape pipeline Genome build: z10 Supplementary files format and content: tab delimited text files for icshape files. First column is the Refseq ID second is transcript length third is RPKM then every column is the nucleotide resolution icSHAPE score. NULL means no confident score. Tab delimited text files for read counts and rpkm files. First column is the Refseq ID second is transcript length third is the unique mapped reads count fourth is the mulitple mapped reads count fifth is the RPKM.", "1 cell NAIN3", "For in vivo treatment cells was incubated with NAI N3 or DMSO at 28.5 C for 5min.", "The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol\u00ae Reagent invitrogen.To enrich polyA RNA the total RNA was subjected to polyA selection. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37\u00b0C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/\u00b5l RiboLock 0.1 U/\u00b5l FastAP Life Technology 2 U/\u00b5l T4 PNKNEB before 3\u2019 adapter ligation with addition of 10 \u00b5l ligation mix 5mM DTT 0.5\u00b5M 5\u2019 adenylated and 3\u2019 blocked linker 3\u2019 bio for unmodified 3\u2019 ddc for modified 0.66 U/\u00b5l T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25\u00b0C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1\u00b5l FastAP 0.2\u00b5l SSB Promega 0.8\u00b5l Ribolock and 1\u00b5l 5\u2019 Deadenylase NEB in 1X NEB buffer 2 NEB at 30\u00b0C for 90 min. And then 1\u00b5l RecJf NEB was added with another incubation at 37\u00b0C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq\uff0c Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer\u2019s protocol.", "Zebrafish wild type strain AB was raised in system water at 28.5\u00b0C under standard conditions.", "strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:0.4hpf", "GSM3409379", "GSM3409379: 1 cell NAIN3; Danio rerio; OTHER", "GSM3409379", null, "1", "The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol\u00ae Reagent invitrogen. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37\u00b0C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/\u00b5l RiboLock 0.1 U/\u00b5l FastAP Life Technology 2 U/\u00b5l T4 PNKNEB before three prime adapter ligation with addition of 10 \u00b5l ligation mix 5mM DTT 0.5\u00b5M five prime adenylated and three prime blocked linker three prime bio for unmodified three prime ddc for modified 0.66 U/\u00b5l T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25\u00b0C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1\u00b5l FastAP 0.2\u00b5l SSB Promega 0.8\u00b5l Ribolock and 1\u00b5l five prime Deadenylase NEB in 1X NEB buffer 2 NEB at 30\u00b0C for 90 min. And then 1\u00b5l RecJf NEB was added with another incubation at 37\u00b0C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq\uff0c Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer's protocol.", "GEO Accession:GSM3409379", "OTHER", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP163087", null, null, "1-cell_NAIN3_rep2.fastq.gz", "fastq", 97797667950.0, 651984453.0, "GSM3409379 r2", "0:150 1:0", "A:28116666638;C:24142558968;G:22646127152;T:22878466770;N:13848422", 150, 0, null, null, 28116666638, 24142558968, 22646127152, 22878466770, 13848422, "SRX4781855", "SRS3862047", "SRA787572", "GEO", "Life Science, Tsinghua University", 1, 0.04751, null, 0.01101, null, 0.93523, null, 0.66405, null, 150, null, "B", null, "usable mapping rate", "illumina", "hiseq_era", "unknown", "poly_a", "unknown", "bulk", "clip", "iclip", null, "China", "2018-10-01", "Zygote", "Embryo", "Embryo Imprecise", "All anatomical structures"], [49605, "SRR7947893", "SRX4781854", "SRS3862045", "SRP163087", "PRJNA494251", "Dynamic landscapes and regulome of mRNA structure during zebrafish early embryogenesis", "GSE120724", "Other", "We resolved the RNA secondary structure during zebrafish early embryogenesis based on in vivo click selective 2' hydroxyl acylation and profiling experiment icSHAPE. We analyzed the RNA structure dynamics among different development stages. Also we studied which factors regulate maternal gene decay by RNA structure switch. Overall design: icSHAPE of 12 samples from zebrafish embryos at six different early development stages and 4 samples from Elavl1a deficient zebrafish embryos at sphere and shield stages including two treated with DMSO as control and two treated with NAIN3 for each stage or condition. RNA seq of 6 samples from zebrafish embryos at 2 4 6 h.p.f. and 4 samples from Elavl1a deficient zebrafish embryos at 4 and 6 h.p.f. including two biological replicates. iCLIP seq of 4 samples from flag elavl1a mRNA injected zebrafish embryos at xxx and 6 h.p.f. including two biological replicates.", null, "pubmed:32423473", null, "1 cell DMSO icSHAPE", "GSM3409378", null, "source name:1 cell DMSO|strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:0.4hpf", "1 cell DMSO icSHAPE", "Library strategy: icSHAPE icSHAPE computational pipeline reference: PubMedID: 26766114 DOI: 10.1038/nprot.2016.011 trimmed three prime adaptor from reads by cutadapt trimmed low quality bases by trimmomatic collapsed duplicated reads as icSHAPE pipeline trimmed the leading 13nt UMI mapping reads to zebrafish reference transcriptomez10 by bowtie2 using the same parameters in icSHAPE pipeline calculate rpkm as icshape pipeline calculate reverse transcription stop sitesRT stop as icshape pipeline RT stop was normalized by using sliding window then icSHAPE score was calculated as icshape pipeline Genome build: z10 Supplementary files format and content: tab delimited text files for icshape files. First column is the Refseq ID second is transcript length third is RPKM then every column is the nucleotide resolution icSHAPE score. NULL means no confident score. Tab delimited text files for read counts and rpkm files. First column is the Refseq ID second is transcript length third is the unique mapped reads count fourth is the mulitple mapped reads count fifth is the RPKM.", "1 cell DMSO", "For in vivo treatment cells was incubated with NAI N3 or DMSO at 28.5 C for 5min.", "The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol\u00ae Reagent invitrogen.To enrich polyA RNA the total RNA was subjected to polyA selection. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37\u00b0C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/\u00b5l RiboLock 0.1 U/\u00b5l FastAP Life Technology 2 U/\u00b5l T4 PNKNEB before 3\u2019 adapter ligation with addition of 10 \u00b5l ligation mix 5mM DTT 0.5\u00b5M 5\u2019 adenylated and 3\u2019 blocked linker 3\u2019 bio for unmodified 3\u2019 ddc for modified 0.66 U/\u00b5l T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25\u00b0C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1\u00b5l FastAP 0.2\u00b5l SSB Promega 0.8\u00b5l Ribolock and 1\u00b5l 5\u2019 Deadenylase NEB in 1X NEB buffer 2 NEB at 30\u00b0C for 90 min. And then 1\u00b5l RecJf NEB was added with another incubation at 37\u00b0C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq\uff0c Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer\u2019s protocol.", "Zebrafish wild type strain AB was raised in system water at 28.5\u00b0C under standard conditions.", "strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:0.4hpf", "GSM3409378", "GSM3409378: 1 cell DMSO; Danio rerio; OTHER", "GSM3409378", null, "1", "The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol\u00ae Reagent invitrogen. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37\u00b0C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/\u00b5l RiboLock 0.1 U/\u00b5l FastAP Life Technology 2 U/\u00b5l T4 PNKNEB before three prime adapter ligation with addition of 10 \u00b5l ligation mix 5mM DTT 0.5\u00b5M five prime adenylated and three prime blocked linker three prime bio for unmodified three prime ddc for modified 0.66 U/\u00b5l T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25\u00b0C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1\u00b5l FastAP 0.2\u00b5l SSB Promega 0.8\u00b5l Ribolock and 1\u00b5l five prime Deadenylase NEB in 1X NEB buffer 2 NEB at 30\u00b0C for 90 min. And then 1\u00b5l RecJf NEB was added with another incubation at 37\u00b0C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq\uff0c Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer's protocol.", "GEO Accession:GSM3409378", "OTHER", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP163087", null, null, "1-cell_DMSO_rep1.fastq.gz", "fastq", 53051740800.0, 353678272.0, "GSM3409378 r1", "0:150 1:0", "A:15070372271;C:13634992007;G:11412296093;T:12928880499;N:5199930", 150, 0, null, null, 15070372271, 13634992007, 11412296093, 12928880499, 5199930, "SRX4781854", "SRS3862045", "SRA787572", "GEO", "Life Science, Tsinghua University", 1, 0.09494, null, 0.00879, null, 0.90319, null, 0.66624, null, 150, null, "B", null, "usable mapping rate", "illumina", "hiseq_era", "unknown", "poly_a", "unknown", "bulk", "clip", "iclip", null, "China", "2018-10-01", "Zygote", "Embryo", "Embryo Imprecise", "All anatomical structures"], [49606, "SRR7947894", "SRX4781854", "SRS3862045", "SRP163087", "PRJNA494251", "Dynamic landscapes and regulome of mRNA structure during zebrafish early embryogenesis", "GSE120724", "Other", "We resolved the RNA secondary structure during zebrafish early embryogenesis based on in vivo click selective 2' hydroxyl acylation and profiling experiment icSHAPE. We analyzed the RNA structure dynamics among different development stages. Also we studied which factors regulate maternal gene decay by RNA structure switch. Overall design: icSHAPE of 12 samples from zebrafish embryos at six different early development stages and 4 samples from Elavl1a deficient zebrafish embryos at sphere and shield stages including two treated with DMSO as control and two treated with NAIN3 for each stage or condition. RNA seq of 6 samples from zebrafish embryos at 2 4 6 h.p.f. and 4 samples from Elavl1a deficient zebrafish embryos at 4 and 6 h.p.f. including two biological replicates. iCLIP seq of 4 samples from flag elavl1a mRNA injected zebrafish embryos at xxx and 6 h.p.f. including two biological replicates.", null, "pubmed:32423473", null, "1 cell DMSO icSHAPE", "GSM3409378", null, "source name:1 cell DMSO|strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:0.4hpf", "1 cell DMSO icSHAPE", "Library strategy: icSHAPE icSHAPE computational pipeline reference: PubMedID: 26766114 DOI: 10.1038/nprot.2016.011 trimmed three prime adaptor from reads by cutadapt trimmed low quality bases by trimmomatic collapsed duplicated reads as icSHAPE pipeline trimmed the leading 13nt UMI mapping reads to zebrafish reference transcriptomez10 by bowtie2 using the same parameters in icSHAPE pipeline calculate rpkm as icshape pipeline calculate reverse transcription stop sitesRT stop as icshape pipeline RT stop was normalized by using sliding window then icSHAPE score was calculated as icshape pipeline Genome build: z10 Supplementary files format and content: tab delimited text files for icshape files. First column is the Refseq ID second is transcript length third is RPKM then every column is the nucleotide resolution icSHAPE score. NULL means no confident score. Tab delimited text files for read counts and rpkm files. First column is the Refseq ID second is transcript length third is the unique mapped reads count fourth is the mulitple mapped reads count fifth is the RPKM.", "1 cell DMSO", "For in vivo treatment cells was incubated with NAI N3 or DMSO at 28.5 C for 5min.", "The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol\u00ae Reagent invitrogen.To enrich polyA RNA the total RNA was subjected to polyA selection. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37\u00b0C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/\u00b5l RiboLock 0.1 U/\u00b5l FastAP Life Technology 2 U/\u00b5l T4 PNKNEB before 3\u2019 adapter ligation with addition of 10 \u00b5l ligation mix 5mM DTT 0.5\u00b5M 5\u2019 adenylated and 3\u2019 blocked linker 3\u2019 bio for unmodified 3\u2019 ddc for modified 0.66 U/\u00b5l T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25\u00b0C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1\u00b5l FastAP 0.2\u00b5l SSB Promega 0.8\u00b5l Ribolock and 1\u00b5l 5\u2019 Deadenylase NEB in 1X NEB buffer 2 NEB at 30\u00b0C for 90 min. And then 1\u00b5l RecJf NEB was added with another incubation at 37\u00b0C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq\uff0c Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer\u2019s protocol.", "Zebrafish wild type strain AB was raised in system water at 28.5\u00b0C under standard conditions.", "strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:0.4hpf", "GSM3409378", "GSM3409378: 1 cell DMSO; Danio rerio; OTHER", "GSM3409378", null, "1", "The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol\u00ae Reagent invitrogen. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37\u00b0C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/\u00b5l RiboLock 0.1 U/\u00b5l FastAP Life Technology 2 U/\u00b5l T4 PNKNEB before three prime adapter ligation with addition of 10 \u00b5l ligation mix 5mM DTT 0.5\u00b5M five prime adenylated and three prime blocked linker three prime bio for unmodified three prime ddc for modified 0.66 U/\u00b5l T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25\u00b0C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1\u00b5l FastAP 0.2\u00b5l SSB Promega 0.8\u00b5l Ribolock and 1\u00b5l five prime Deadenylase NEB in 1X NEB buffer 2 NEB at 30\u00b0C for 90 min. And then 1\u00b5l RecJf NEB was added with another incubation at 37\u00b0C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq\uff0c Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer's protocol.", "GEO Accession:GSM3409378", "OTHER", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP163087", null, null, "1-cell_DMSO_rep2.fastq.gz", "fastq", 63247728900.0, 421651526.0, "GSM3409378 r2", "0:150 1:0", "A:18082840284;C:14901926799;G:14919348528;T:15334620404;N:8992885", 150, 0, null, null, 18082840284, 14901926799, 14919348528, 15334620404, 8992885, "SRX4781854", "SRS3862045", "SRA787572", "GEO", "Life Science, Tsinghua University", 1, 0.07037, null, 0.00751, null, 0.91662, null, 0.68677, null, 150, null, "B", null, "usable mapping rate", "illumina", "hiseq_era", "unknown", "poly_a", "unknown", "bulk", "clip", "iclip", null, "China", "2018-10-01", "Zygote", "Embryo", "Embryo Imprecise", "All anatomical structures"], [49607, "SRR7947891", "SRX4781853", "SRS3862046", "SRP163087", "PRJNA494251", "Dynamic landscapes and regulome of mRNA structure during zebrafish early embryogenesis", "GSE120724", "Other", "We resolved the RNA secondary structure during zebrafish early embryogenesis based on in vivo click selective 2' hydroxyl acylation and profiling experiment icSHAPE. We analyzed the RNA structure dynamics among different development stages. Also we studied which factors regulate maternal gene decay by RNA structure switch. Overall design: icSHAPE of 12 samples from zebrafish embryos at six different early development stages and 4 samples from Elavl1a deficient zebrafish embryos at sphere and shield stages including two treated with DMSO as control and two treated with NAIN3 for each stage or condition. RNA seq of 6 samples from zebrafish embryos at 2 4 6 h.p.f. and 4 samples from Elavl1a deficient zebrafish embryos at 4 and 6 h.p.f. including two biological replicates. iCLIP seq of 4 samples from flag elavl1a mRNA injected zebrafish embryos at xxx and 6 h.p.f. including two biological replicates.", null, "pubmed:32423473", null, "fertilized egg NAIN3 icSHAPE", "GSM3409377", null, "source name:fertilized egg NAIN3|strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:0hpf", "fertilized egg NAIN3 icSHAPE", "Library strategy: icSHAPE icSHAPE computational pipeline reference: PubMedID: 26766114 DOI: 10.1038/nprot.2016.011 trimmed three prime adaptor from reads by cutadapt trimmed low quality bases by trimmomatic collapsed duplicated reads as icSHAPE pipeline trimmed the leading 13nt UMI mapping reads to zebrafish reference transcriptomez10 by bowtie2 using the same parameters in icSHAPE pipeline calculate rpkm as icshape pipeline calculate reverse transcription stop sitesRT stop as icshape pipeline RT stop was normalized by using sliding window then icSHAPE score was calculated as icshape pipeline Genome build: z10 Supplementary files format and content: tab delimited text files for icshape files. First column is the Refseq ID second is transcript length third is RPKM then every column is the nucleotide resolution icSHAPE score. NULL means no confident score. Tab delimited text files for read counts and rpkm files. First column is the Refseq ID second is transcript length third is the unique mapped reads count fourth is the mulitple mapped reads count fifth is the RPKM.", "fertilized egg NAIN3", "For in vivo treatment cells was incubated with NAI N3 or DMSO at 28.5 C for 5min.", "The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol\u00ae Reagent invitrogen.To enrich polyA RNA the total RNA was subjected to polyA selection. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37\u00b0C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/\u00b5l RiboLock 0.1 U/\u00b5l FastAP Life Technology 2 U/\u00b5l T4 PNKNEB before 3\u2019 adapter ligation with addition of 10 \u00b5l ligation mix 5mM DTT 0.5\u00b5M 5\u2019 adenylated and 3\u2019 blocked linker 3\u2019 bio for unmodified 3\u2019 ddc for modified 0.66 U/\u00b5l T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25\u00b0C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1\u00b5l FastAP 0.2\u00b5l SSB Promega 0.8\u00b5l Ribolock and 1\u00b5l 5\u2019 Deadenylase NEB in 1X NEB buffer 2 NEB at 30\u00b0C for 90 min. And then 1\u00b5l RecJf NEB was added with another incubation at 37\u00b0C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq\uff0c Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer\u2019s protocol.", "Zebrafish wild type strain AB was raised in system water at 28.5\u00b0C under standard conditions.", "strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:0hpf", "GSM3409377", "GSM3409377: fertilized egg NAIN3; Danio rerio; OTHER", "GSM3409377", null, "1", "The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol\u00ae Reagent invitrogen. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37\u00b0C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/\u00b5l RiboLock 0.1 U/\u00b5l FastAP Life Technology 2 U/\u00b5l T4 PNKNEB before three prime adapter ligation with addition of 10 \u00b5l ligation mix 5mM DTT 0.5\u00b5M five prime adenylated and three prime blocked linker three prime bio for unmodified three prime ddc for modified 0.66 U/\u00b5l T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25\u00b0C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1\u00b5l FastAP 0.2\u00b5l SSB Promega 0.8\u00b5l Ribolock and 1\u00b5l five prime Deadenylase NEB in 1X NEB buffer 2 NEB at 30\u00b0C for 90 min. And then 1\u00b5l RecJf NEB was added with another incubation at 37\u00b0C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq\uff0c Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer's protocol.", "GEO Accession:GSM3409377", "OTHER", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP163087", null, null, "fertilized_egg_NAIN3_rep1.fastq.gz", "fastq", 142104356700.0, 947362378.0, "GSM3409377 r1", "0:150 1:0", "A:43254172321;C:33297470356;G:29006285816;T:36529804597;N:16623610", 150, 0, null, null, 43254172321, 33297470356, 29006285816, 36529804597, 16623610, "SRX4781853", "SRS3862046", "SRA787572", "GEO", "Life Science, Tsinghua University", 1, 0.02945, null, 0.00559, null, 0.95875, null, 0.73322, null, 150, null, "B", null, "usable mapping rate", "illumina", "hiseq_era", "unknown", "poly_a", "unknown", "bulk", "clip", "iclip", null, "China", "2018-10-01", "Zygote", "Embryo", "Embryo Imprecise", "All anatomical structures"], [49608, "SRR7947892", "SRX4781853", "SRS3862046", "SRP163087", "PRJNA494251", "Dynamic landscapes and regulome of mRNA structure during zebrafish early embryogenesis", "GSE120724", "Other", "We resolved the RNA secondary structure during zebrafish early embryogenesis based on in vivo click selective 2' hydroxyl acylation and profiling experiment icSHAPE. We analyzed the RNA structure dynamics among different development stages. Also we studied which factors regulate maternal gene decay by RNA structure switch. Overall design: icSHAPE of 12 samples from zebrafish embryos at six different early development stages and 4 samples from Elavl1a deficient zebrafish embryos at sphere and shield stages including two treated with DMSO as control and two treated with NAIN3 for each stage or condition. RNA seq of 6 samples from zebrafish embryos at 2 4 6 h.p.f. and 4 samples from Elavl1a deficient zebrafish embryos at 4 and 6 h.p.f. including two biological replicates. iCLIP seq of 4 samples from flag elavl1a mRNA injected zebrafish embryos at xxx and 6 h.p.f. including two biological replicates.", null, "pubmed:32423473", null, "fertilized egg NAIN3 icSHAPE", "GSM3409377", null, "source name:fertilized egg NAIN3|strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:0hpf", "fertilized egg NAIN3 icSHAPE", "Library strategy: icSHAPE icSHAPE computational pipeline reference: PubMedID: 26766114 DOI: 10.1038/nprot.2016.011 trimmed three prime adaptor from reads by cutadapt trimmed low quality bases by trimmomatic collapsed duplicated reads as icSHAPE pipeline trimmed the leading 13nt UMI mapping reads to zebrafish reference transcriptomez10 by bowtie2 using the same parameters in icSHAPE pipeline calculate rpkm as icshape pipeline calculate reverse transcription stop sitesRT stop as icshape pipeline RT stop was normalized by using sliding window then icSHAPE score was calculated as icshape pipeline Genome build: z10 Supplementary files format and content: tab delimited text files for icshape files. First column is the Refseq ID second is transcript length third is RPKM then every column is the nucleotide resolution icSHAPE score. NULL means no confident score. Tab delimited text files for read counts and rpkm files. First column is the Refseq ID second is transcript length third is the unique mapped reads count fourth is the mulitple mapped reads count fifth is the RPKM.", "fertilized egg NAIN3", "For in vivo treatment cells was incubated with NAI N3 or DMSO at 28.5 C for 5min.", "The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol\u00ae Reagent invitrogen.To enrich polyA RNA the total RNA was subjected to polyA selection. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37\u00b0C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/\u00b5l RiboLock 0.1 U/\u00b5l FastAP Life Technology 2 U/\u00b5l T4 PNKNEB before 3\u2019 adapter ligation with addition of 10 \u00b5l ligation mix 5mM DTT 0.5\u00b5M 5\u2019 adenylated and 3\u2019 blocked linker 3\u2019 bio for unmodified 3\u2019 ddc for modified 0.66 U/\u00b5l T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25\u00b0C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1\u00b5l FastAP 0.2\u00b5l SSB Promega 0.8\u00b5l Ribolock and 1\u00b5l 5\u2019 Deadenylase NEB in 1X NEB buffer 2 NEB at 30\u00b0C for 90 min. And then 1\u00b5l RecJf NEB was added with another incubation at 37\u00b0C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq\uff0c Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer\u2019s protocol.", "Zebrafish wild type strain AB was raised in system water at 28.5\u00b0C under standard conditions.", "strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:0hpf", "GSM3409377", "GSM3409377: fertilized egg NAIN3; Danio rerio; OTHER", "GSM3409377", null, "1", "The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol\u00ae Reagent invitrogen. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37\u00b0C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/\u00b5l RiboLock 0.1 U/\u00b5l FastAP Life Technology 2 U/\u00b5l T4 PNKNEB before three prime adapter ligation with addition of 10 \u00b5l ligation mix 5mM DTT 0.5\u00b5M five prime adenylated and three prime blocked linker three prime bio for unmodified three prime ddc for modified 0.66 U/\u00b5l T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25\u00b0C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1\u00b5l FastAP 0.2\u00b5l SSB Promega 0.8\u00b5l Ribolock and 1\u00b5l five prime Deadenylase NEB in 1X NEB buffer 2 NEB at 30\u00b0C for 90 min. And then 1\u00b5l RecJf NEB was added with another incubation at 37\u00b0C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq\uff0c Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer's protocol.", "GEO Accession:GSM3409377", "OTHER", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP163087", null, null, "fertilized_egg_NAIN3_rep2.fastq.gz", "fastq", 116798258100.0, 778655054.0, "GSM3409377 r2", "0:150 1:0", "A:35299190309;C:27819081594;G:24802574513;T:28860009513;N:17402171", 150, 0, null, null, 35299190309, 27819081594, 24802574513, 28860009513, 17402171, "SRX4781853", "SRS3862046", "SRA787572", "GEO", "Life Science, Tsinghua University", 1, 0.04759, null, 0.01267, null, 0.94483, null, 0.69919, null, 150, null, "B", null, "usable mapping rate", "illumina", "hiseq_era", "unknown", "poly_a", "unknown", "bulk", "clip", "iclip", null, "China", "2018-10-01", "Zygote", "Embryo", "Embryo Imprecise", "All anatomical structures"], [49609, "SRR7947889", "SRX4781852", "SRS3862044", "SRP163087", "PRJNA494251", "Dynamic landscapes and regulome of mRNA structure during zebrafish early embryogenesis", "GSE120724", "Other", "We resolved the RNA secondary structure during zebrafish early embryogenesis based on in vivo click selective 2' hydroxyl acylation and profiling experiment icSHAPE. We analyzed the RNA structure dynamics among different development stages. Also we studied which factors regulate maternal gene decay by RNA structure switch. Overall design: icSHAPE of 12 samples from zebrafish embryos at six different early development stages and 4 samples from Elavl1a deficient zebrafish embryos at sphere and shield stages including two treated with DMSO as control and two treated with NAIN3 for each stage or condition. RNA seq of 6 samples from zebrafish embryos at 2 4 6 h.p.f. and 4 samples from Elavl1a deficient zebrafish embryos at 4 and 6 h.p.f. including two biological replicates. iCLIP seq of 4 samples from flag elavl1a mRNA injected zebrafish embryos at xxx and 6 h.p.f. including two biological replicates.", null, "pubmed:32423473", null, "fertilized egg DMSO icSHAPE", "GSM3409376", null, "source name:fertilized egg DMSO|strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:0hpf", "fertilized egg DMSO icSHAPE", "Library strategy: icSHAPE icSHAPE computational pipeline reference: PubMedID: 26766114 DOI: 10.1038/nprot.2016.011 trimmed three prime adaptor from reads by cutadapt trimmed low quality bases by trimmomatic collapsed duplicated reads as icSHAPE pipeline trimmed the leading 13nt UMI mapping reads to zebrafish reference transcriptomez10 by bowtie2 using the same parameters in icSHAPE pipeline calculate rpkm as icshape pipeline calculate reverse transcription stop sitesRT stop as icshape pipeline RT stop was normalized by using sliding window then icSHAPE score was calculated as icshape pipeline Genome build: z10 Supplementary files format and content: tab delimited text files for icshape files. First column is the Refseq ID second is transcript length third is RPKM then every column is the nucleotide resolution icSHAPE score. NULL means no confident score. Tab delimited text files for read counts and rpkm files. First column is the Refseq ID second is transcript length third is the unique mapped reads count fourth is the mulitple mapped reads count fifth is the RPKM.", "fertilized egg DMSO", "For in vivo treatment cells was incubated with NAI N3 or DMSO at 28.5 C for 5min.", "The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol\u00ae Reagent invitrogen.To enrich polyA RNA the total RNA was subjected to polyA selection. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37\u00b0C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/\u00b5l RiboLock 0.1 U/\u00b5l FastAP Life Technology 2 U/\u00b5l T4 PNKNEB before 3\u2019 adapter ligation with addition of 10 \u00b5l ligation mix 5mM DTT 0.5\u00b5M 5\u2019 adenylated and 3\u2019 blocked linker 3\u2019 bio for unmodified 3\u2019 ddc for modified 0.66 U/\u00b5l T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25\u00b0C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1\u00b5l FastAP 0.2\u00b5l SSB Promega 0.8\u00b5l Ribolock and 1\u00b5l 5\u2019 Deadenylase NEB in 1X NEB buffer 2 NEB at 30\u00b0C for 90 min. And then 1\u00b5l RecJf NEB was added with another incubation at 37\u00b0C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq\uff0c Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer\u2019s protocol.", "Zebrafish wild type strain AB was raised in system water at 28.5\u00b0C under standard conditions.", "strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:0hpf", "GSM3409376", "GSM3409376: fertilized egg DMSO; Danio rerio; OTHER", "GSM3409376", null, "1", "The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol\u00ae Reagent invitrogen. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37\u00b0C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/\u00b5l RiboLock 0.1 U/\u00b5l FastAP Life Technology 2 U/\u00b5l T4 PNKNEB before three prime adapter ligation with addition of 10 \u00b5l ligation mix 5mM DTT 0.5\u00b5M five prime adenylated and three prime blocked linker three prime bio for unmodified three prime ddc for modified 0.66 U/\u00b5l T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25\u00b0C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1\u00b5l FastAP 0.2\u00b5l SSB Promega 0.8\u00b5l Ribolock and 1\u00b5l five prime Deadenylase NEB in 1X NEB buffer 2 NEB at 30\u00b0C for 90 min. And then 1\u00b5l RecJf NEB was added with another incubation at 37\u00b0C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq\uff0c Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer's protocol.", "GEO Accession:GSM3409376", "OTHER", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP163087", null, null, "fertilized_egg_DMSO_rep1.fastq.gz", "fastq", 34356249600.0, 229041664.0, "GSM3409376 r1", "0:150 1:0", "A:9379426193;C:8993497696;G:7413604416;T:8565620110;N:4101185", 150, 0, null, null, 9379426193, 8993497696, 7413604416, 8565620110, 4101185, "SRX4781852", "SRS3862044", "SRA787572", "GEO", "Life Science, Tsinghua University", 1, 0.03911, null, 0.0035, null, 0.9441, null, 0.74394, null, 150, null, "B", null, "usable mapping rate", "illumina", "hiseq_era", "unknown", "poly_a", "unknown", "bulk", "clip", "iclip", null, "China", "2018-10-01", "Zygote", "Embryo", "Embryo Imprecise", "All anatomical structures"], [49610, "SRR7947890", "SRX4781852", "SRS3862044", "SRP163087", "PRJNA494251", "Dynamic landscapes and regulome of mRNA structure during zebrafish early embryogenesis", "GSE120724", "Other", "We resolved the RNA secondary structure during zebrafish early embryogenesis based on in vivo click selective 2' hydroxyl acylation and profiling experiment icSHAPE. We analyzed the RNA structure dynamics among different development stages. Also we studied which factors regulate maternal gene decay by RNA structure switch. Overall design: icSHAPE of 12 samples from zebrafish embryos at six different early development stages and 4 samples from Elavl1a deficient zebrafish embryos at sphere and shield stages including two treated with DMSO as control and two treated with NAIN3 for each stage or condition. RNA seq of 6 samples from zebrafish embryos at 2 4 6 h.p.f. and 4 samples from Elavl1a deficient zebrafish embryos at 4 and 6 h.p.f. including two biological replicates. iCLIP seq of 4 samples from flag elavl1a mRNA injected zebrafish embryos at xxx and 6 h.p.f. including two biological replicates.", null, "pubmed:32423473", null, "fertilized egg DMSO icSHAPE", "GSM3409376", null, "source name:fertilized egg DMSO|strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:0hpf", "fertilized egg DMSO icSHAPE", "Library strategy: icSHAPE icSHAPE computational pipeline reference: PubMedID: 26766114 DOI: 10.1038/nprot.2016.011 trimmed three prime adaptor from reads by cutadapt trimmed low quality bases by trimmomatic collapsed duplicated reads as icSHAPE pipeline trimmed the leading 13nt UMI mapping reads to zebrafish reference transcriptomez10 by bowtie2 using the same parameters in icSHAPE pipeline calculate rpkm as icshape pipeline calculate reverse transcription stop sitesRT stop as icshape pipeline RT stop was normalized by using sliding window then icSHAPE score was calculated as icshape pipeline Genome build: z10 Supplementary files format and content: tab delimited text files for icshape files. First column is the Refseq ID second is transcript length third is RPKM then every column is the nucleotide resolution icSHAPE score. NULL means no confident score. Tab delimited text files for read counts and rpkm files. First column is the Refseq ID second is transcript length third is the unique mapped reads count fourth is the mulitple mapped reads count fifth is the RPKM.", "fertilized egg DMSO", "For in vivo treatment cells was incubated with NAI N3 or DMSO at 28.5 C for 5min.", "The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol\u00ae Reagent invitrogen.To enrich polyA RNA the total RNA was subjected to polyA selection. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37\u00b0C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/\u00b5l RiboLock 0.1 U/\u00b5l FastAP Life Technology 2 U/\u00b5l T4 PNKNEB before 3\u2019 adapter ligation with addition of 10 \u00b5l ligation mix 5mM DTT 0.5\u00b5M 5\u2019 adenylated and 3\u2019 blocked linker 3\u2019 bio for unmodified 3\u2019 ddc for modified 0.66 U/\u00b5l T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25\u00b0C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1\u00b5l FastAP 0.2\u00b5l SSB Promega 0.8\u00b5l Ribolock and 1\u00b5l 5\u2019 Deadenylase NEB in 1X NEB buffer 2 NEB at 30\u00b0C for 90 min. And then 1\u00b5l RecJf NEB was added with another incubation at 37\u00b0C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq\uff0c Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer\u2019s protocol.", "Zebrafish wild type strain AB was raised in system water at 28.5\u00b0C under standard conditions.", "strain:AB|tissue:zebrafish embryos|genotype/variation:normal zebrafish embryos cells|age:0hpf", "GSM3409376", "GSM3409376: fertilized egg DMSO; Danio rerio; OTHER", "GSM3409376", null, "1", "The RNA or modified RNA was extracted or cleaned up and eluted in RNase free water by using TRIzol\u00ae Reagent invitrogen. For icSHAPE the in vivo modified RNA and unmodified RNA were biotinylated by copper free click reaction followed by fragmentation. The fragmented RNA was end repaired by incubation at 37\u00b0C for 1 hours with the following mix 70 mM Tris 7.0 18mM MgCl2 5mM DTT 4 U/\u00b5l RiboLock 0.1 U/\u00b5l FastAP Life Technology 2 U/\u00b5l T4 PNKNEB before three prime adapter ligation with addition of 10 \u00b5l ligation mix 5mM DTT 0.5\u00b5M five prime adenylated and three prime blocked linker three prime bio for unmodified three prime ddc for modified 0.66 U/\u00b5l T4 RNA ligase NEB M0437M 15% PEG8000 1X RNA ligase buffer and incubation at 25\u00b0C for extra 3 hours. Then the excess adaptor was removed as described below. The purified RNA was incubated in the mix of 1\u00b5l FastAP 0.2\u00b5l SSB Promega 0.8\u00b5l Ribolock and 1\u00b5l five prime Deadenylase NEB in 1X NEB buffer 2 NEB at 30\u00b0C for 90 min. And then 1\u00b5l RecJf NEB was added with another incubation at 37\u00b0C for 1 hour. The following procedures including reverse transcription biotin streptavadin enrichment size selection of cDNA circularization and PCR amplification were the same as described in the standard protocol. For RNA seq\uff0c Fragmented mRNA was used for library construction using the KAPA Stranded mRNA Seq Kit KAPA according to manufacturer's protocol.", "GEO Accession:GSM3409376", "OTHER", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina HiSeq 2500", null, "SRP163087", null, null, "fertilized_egg_DMSO_rep2.fastq.gz", "fastq", 44580988350.0, 297206589.0, "GSM3409376 r2", "0:150 1:0", "A:12959202147;C:11011600906;G:9852124926;T:10754108697;N:3951674", 150, 0, null, null, 12959202147, 11011600906, 9852124926, 10754108697, 3951674, "SRX4781852", "SRS3862044", "SRA787572", "GEO", "Life Science, Tsinghua University", 1, 0.05584, null, 0.00851, null, 0.93604, null, 0.7312, null, 150, null, "B", null, "usable mapping rate", "illumina", "hiseq_era", "unknown", "poly_a", "unknown", "bulk", "clip", "iclip", null, "China", "2018-10-01", "Zygote", "Embryo", "Embryo Imprecise", "All anatomical structures"], [59512, "SRR11924307", "SRX8469983", "SRS6770634", "SRP265951", "PRJNA637293", "The shift from early to late types of ribosomes in zebrafish development involves changes at a subset of rRNA 2' O Me sites", "GSE151797", "Other", "A sequencing based profiling method RiboMeth seq for ribose methylations was used to study methylation patterns during Zebrafish Danio rerio development Overall design: All samples were analyzed in biological triplicates except for adult tail trunk that was in duplicate.", null, "pubmed:32912962", null, "Unf egg 3", "GSM4591051", null, "source name:unfertilized egg|tissue:unfertilized egg|rna fraction:size fractionated 20 40 nt whole cell RNA", "Unf egg 3", "Library strategy: RiboMeth seq Barcode separation using python script Adaptor trimming using Cutadapt v. 2.0 Mapping to rRNA reference sequence using Bowtie2 v. 2.3.4.1 Counting read ends and calculating RiboMeth seq scores using python scripts The output FASTA files from small RNA seq were merged and used as the basis of the SNORD search and rRNA interaction prediction. Initially SNORDs were identified by running the merged FASTA file through snoScan Schattner et al. 2005 against zebrafish early and late rRNA reference sequences Locati et al. 2017. Genome build: early and late zebrafish rRNA locati et al. The reference sequences are available in the FASTA file on the series record. Supplementary files format and content: MS Excel file contains five prime and three prime read count and calculated RiboMeth seq score at all positions in the rRNA sequence.", "unfertilized egg", null, "Tissues were homogenized and whole cell RNA was extracted using Qiazol Qiagen according to the manufacturer. RiboMeth seq: 5 10 ug of RNA was partially degraded by alkaline at denaturing temperatures. The size fraction 20 40 nt was purified on gels and linkers added using a system relying on a modified Arabidopsis tRNA ligase joining 2' three prime cyclic phosphate and five prime phosphate ends. The library fragments were then sequenced on the Ion Proton platform. See Birkedal U Christensen Dalsgaard M Krogh N Sabarinathan R Gorodkin J Nielsen H. Profiling of ribose methylations in RNA by high throughput sequencing. Angewandte Chemie. 2015;542:451 5 for detailed description", null, "tissue:unfertilized egg|rna fraction:size fractionated 20 40 nt whole cell RNA", "GSM4591051", "GSM4591051: Unf egg 3; Danio rerio; OTHER", "GSM4591051", null, "1", "Tissues were homogenized and whole cell RNA was extracted using Qiazol Qiagen according to the manufacturer. RiboMeth seq: 5 10 ug of RNA was partially degraded by alkaline at denaturing temperatures. The size fraction 20 40 nt was purified on gels and linkers added using a system relying on a modified Arabidopsis tRNA ligase joining 2' three prime cyclic phosphate and five prime phosphate ends. The library fragments were then sequenced on the Ion Proton platform. See Birkedal U Christensen Dalsgaard M Krogh N Sabarinathan R Gorodkin J Nielsen H. Profiling of ribose methylations in RNA by high throughput sequencing. Angewandte Chemie. 2015;542:451 5 for detailed description", "GEO Accession:GSM4591051", "OTHER", "TRANSCRIPTOMIC", "other", "SINGLE", "ION_TORRENT", "Ion Torrent Proton", null, "SRP265951", null, "intentional duplicate", "GSE151797_Reference_sequence.fa Unf_egg_3.bam", "bam bam", 149835556.0, 5777340.0, "GSM4591051 r1", "0:25.94", "A:27873539;C:50837956;G:37710769;T:33413292;N:0", 25, null, null, null, 27873539, 50837956, 37710769, 33413292, 0, "SRX8469983", "SRS6770634", "SRA1083099", "GEO", "RNA Group - Prof. Henrik Nielsen, Department of Cellular and Molecular Medicine, University of Copenhagen", 1, 0.74561, null, 0.22616, null, 0.89441, null, 0.76897, null, 34, null, "B", null, "usable mapping rate", "ion_torrent", "ion_torrent", "5prime", "small_rna", "unknown", "bulk", "unknown", "unknown", null, "Denmark", "2020-06-04", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [59513, "SRR11924305", "SRX8469982", "SRS6770633", "SRP265951", "PRJNA637293", "The shift from early to late types of ribosomes in zebrafish development involves changes at a subset of rRNA 2' O Me sites", "GSE151797", "Other", "A sequencing based profiling method RiboMeth seq for ribose methylations was used to study methylation patterns during Zebrafish Danio rerio development Overall design: All samples were analyzed in biological triplicates except for adult tail trunk that was in duplicate.", null, "pubmed:32912962", null, "Unf egg 2", "GSM4591050", null, "source name:unfertilized egg|tissue:unfertilized egg|rna fraction:size fractionated 20 40 nt whole cell RNA", "Unf egg 2", "Library strategy: RiboMeth seq Barcode separation using python script Adaptor trimming using Cutadapt v. 2.0 Mapping to rRNA reference sequence using Bowtie2 v. 2.3.4.1 Counting read ends and calculating RiboMeth seq scores using python scripts The output FASTA files from small RNA seq were merged and used as the basis of the SNORD search and rRNA interaction prediction. Initially SNORDs were identified by running the merged FASTA file through snoScan Schattner et al. 2005 against zebrafish early and late rRNA reference sequences Locati et al. 2017. Genome build: early and late zebrafish rRNA locati et al. The reference sequences are available in the FASTA file on the series record. Supplementary files format and content: MS Excel file contains five prime and three prime read count and calculated RiboMeth seq score at all positions in the rRNA sequence.", "unfertilized egg", null, "Tissues were homogenized and whole cell RNA was extracted using Qiazol Qiagen according to the manufacturer. RiboMeth seq: 5 10 ug of RNA was partially degraded by alkaline at denaturing temperatures. The size fraction 20 40 nt was purified on gels and linkers added using a system relying on a modified Arabidopsis tRNA ligase joining 2' three prime cyclic phosphate and five prime phosphate ends. The library fragments were then sequenced on the Ion Proton platform. See Birkedal U Christensen Dalsgaard M Krogh N Sabarinathan R Gorodkin J Nielsen H. Profiling of ribose methylations in RNA by high throughput sequencing. Angewandte Chemie. 2015;542:451 5 for detailed description", null, "tissue:unfertilized egg|rna fraction:size fractionated 20 40 nt whole cell RNA", "GSM4591050", "GSM4591050: Unf egg 2; Danio rerio; OTHER", "GSM4591050", null, "1", "Tissues were homogenized and whole cell RNA was extracted using Qiazol Qiagen according to the manufacturer. RiboMeth seq: 5 10 ug of RNA was partially degraded by alkaline at denaturing temperatures. The size fraction 20 40 nt was purified on gels and linkers added using a system relying on a modified Arabidopsis tRNA ligase joining 2' three prime cyclic phosphate and five prime phosphate ends. The library fragments were then sequenced on the Ion Proton platform. See Birkedal U Christensen Dalsgaard M Krogh N Sabarinathan R Gorodkin J Nielsen H. Profiling of ribose methylations in RNA by high throughput sequencing. Angewandte Chemie. 2015;542:451 5 for detailed description", "GEO Accession:GSM4591050", "OTHER", "TRANSCRIPTOMIC", "other", "SINGLE", "ION_TORRENT", "Ion Torrent Proton", null, "SRP265951", null, "intentional duplicate", "GSE151797_Reference_sequence.fa Unf_egg_2.bam", "bam bam", 235014832.0, 8915586.0, "GSM4591050 r1", "0:26.36", "A:43825969;C:79775479;G:60531219;T:50882165;N:0", 26, null, null, null, 43825969, 79775479, 60531219, 50882165, 0, "SRX8469982", "SRS6770633", "SRA1083099", "GEO", "RNA Group - Prof. Henrik Nielsen, Department of Cellular and Molecular Medicine, University of Copenhagen", 1, 0.78414, null, 0.23044, null, 0.90352, null, 0.76934, null, 40, null, "B", null, "usable mapping rate", "ion_torrent", "ion_torrent", "5prime", "small_rna", "unknown", "bulk", "unknown", "unknown", null, "Denmark", "2020-06-04", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [59514, "SRR11924304", "SRX8469981", "SRS6770632", "SRP265951", "PRJNA637293", "The shift from early to late types of ribosomes in zebrafish development involves changes at a subset of rRNA 2' O Me sites", "GSE151797", "Other", "A sequencing based profiling method RiboMeth seq for ribose methylations was used to study methylation patterns during Zebrafish Danio rerio development Overall design: All samples were analyzed in biological triplicates except for adult tail trunk that was in duplicate.", null, "pubmed:32912962", null, "Unf egg 1", "GSM4591049", null, "source name:unfertilized egg|tissue:unfertilized egg|rna fraction:size fractionated 20 40 nt whole cell RNA", "Unf egg 1", "Library strategy: RiboMeth seq Barcode separation using python script Adaptor trimming using Cutadapt v. 2.0 Mapping to rRNA reference sequence using Bowtie2 v. 2.3.4.1 Counting read ends and calculating RiboMeth seq scores using python scripts The output FASTA files from small RNA seq were merged and used as the basis of the SNORD search and rRNA interaction prediction. Initially SNORDs were identified by running the merged FASTA file through snoScan Schattner et al. 2005 against zebrafish early and late rRNA reference sequences Locati et al. 2017. Genome build: early and late zebrafish rRNA locati et al. The reference sequences are available in the FASTA file on the series record. Supplementary files format and content: MS Excel file contains five prime and three prime read count and calculated RiboMeth seq score at all positions in the rRNA sequence.", "unfertilized egg", null, "Tissues were homogenized and whole cell RNA was extracted using Qiazol Qiagen according to the manufacturer. RiboMeth seq: 5 10 ug of RNA was partially degraded by alkaline at denaturing temperatures. The size fraction 20 40 nt was purified on gels and linkers added using a system relying on a modified Arabidopsis tRNA ligase joining 2' three prime cyclic phosphate and five prime phosphate ends. The library fragments were then sequenced on the Ion Proton platform. See Birkedal U Christensen Dalsgaard M Krogh N Sabarinathan R Gorodkin J Nielsen H. Profiling of ribose methylations in RNA by high throughput sequencing. Angewandte Chemie. 2015;542:451 5 for detailed description", null, "tissue:unfertilized egg|rna fraction:size fractionated 20 40 nt whole cell RNA", "GSM4591049", "GSM4591049: Unf egg 1; Danio rerio; OTHER", "GSM4591049", null, "1", "Tissues were homogenized and whole cell RNA was extracted using Qiazol Qiagen according to the manufacturer. RiboMeth seq: 5 10 ug of RNA was partially degraded by alkaline at denaturing temperatures. The size fraction 20 40 nt was purified on gels and linkers added using a system relying on a modified Arabidopsis tRNA ligase joining 2' three prime cyclic phosphate and five prime phosphate ends. The library fragments were then sequenced on the Ion Proton platform. See Birkedal U Christensen Dalsgaard M Krogh N Sabarinathan R Gorodkin J Nielsen H. Profiling of ribose methylations in RNA by high throughput sequencing. Angewandte Chemie. 2015;542:451 5 for detailed description", "GEO Accession:GSM4591049", "OTHER", "TRANSCRIPTOMIC", "other", "SINGLE", "ION_TORRENT", "Ion Torrent Proton", null, "SRP265951", null, "intentional duplicate", "GSE151797_Reference_sequence.fa Unf_egg_1.bam", "bam bam", 245529952.0, 9182415.0, "GSM4591049 r1", "0:26.74", "A:46557080;C:80893860;G:66050448;T:52028564;N:0", 26, null, null, null, 46557080, 80893860, 66050448, 52028564, 0, "SRX8469981", "SRS6770632", "SRA1083099", "GEO", "RNA Group - Prof. Henrik Nielsen, Department of Cellular and Molecular Medicine, University of Copenhagen", 1, 0.87423, null, 0.27943, null, 0.91721, null, 0.76816, null, 37, null, "B", null, "usable mapping rate", "ion_torrent", "ion_torrent", "5prime", "small_rna", "unknown", "bulk", "unknown", "unknown", null, "Denmark", "2020-06-04", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [66194, "SRR16096233", "SRX12382357", "SRS10353019", "SRP339082", "PRJNA766854", "RNA m6A methylome from zebrafish zygotes mouse liver polyA selected RNAs mouse embryonic stem cells spike in RNA controls and single mouse oocytes and embryos", "GSE184893", "Other", "N6 methyladenosine m6A is a prevalent RNA modification that has key roles in distinct biological processes including meiosis and early embryo development. Despite advances in methodology for m6A mapping the applicability of current methods is limited by the need for large amounts of input material. We develop a sensitive picogram scale m6A RNA immunoprecipitation and sequencing picoMeRIP seq method also suitable for single cell scMeRIP seq. We validate the down scaling of the method by profiling m6A from picogram amounts of mouse liver polyA selected RNA mouse embryonic stem cells and zebrafish zygotes. We provide proof of principle of m6A profiling in single mouse oocytes and preimplantation embryos. Our work opens a new avenue for studying m6A in single cells and scarce cell types in a transcriptome wide manner. Overall design: Transcriptome wide m6A maps were generated from zebrafish and mouse samples as below. 11 zebrafish zygote samples: 3 biological replicates for 10 zygotes 3 biological replicates for 5 zygotes and 5 biological replicates for single zygote. 6 mouse liver polyA selected RNA samples: 10 ng 1 ng and 100 pg; 2 biological replicates for each amount. 9 mouse embryonic stem cell R1 cell line from ATCC SCRC 1011 samples: 1000 cells 100 cells and 10 cells; 3 biological replicates for each amount. 3 spike in RNA control add 0.001 fmol Gaussia luciferase/GLuc with m6A modification and 0.001 fmol Cypridina luciferase/CLuc without xxx modification into the 12 ng polyA selected RNAs related samples: 12 ng polyA selected RNA from mouse liver; 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from wild type/WT mouse embryonic stem cells from Geula S. et al. Science 2015; and 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from Mettl3 knock out/KO mouse embryonic stem cells from Geula S. et al. Science 2015. 12 single mouse oocyte and embryo samples: oocytes at germinal vesicle GV and metaphase II MII stages and embryos at zygote 2 cell 8 cell and blastocyst stages; 2 biological replicates per developmental stage. The pooled Input samples for GV oocyte GSM5746922 zygote GSM5746930 2 cell GSM5746911 and 8 cell GSM5746914 stages can be found in GSE192440.", null, "pubmed:37349523", null, "ZF zygote IP 1cell Rep5", "GSM5599725", null, "tissue:ZF zygote IP 1cell|developmental stage:Zygote|molecule subtype:rRNA free RNA", "ZF zygote IP 1cell Rep5", "Quality of raw reads was assessed using FastQC v0.11.8 and sequencing adapters were trimmed using Cutadapt v1.8.1 if clear adapter signal was detected at the three primeend of the reads. There is no adapter removal for zebrafish zygote samples GSM5599713 GSM5599714 GSM5599715 GSM5599716 GSM5599717 GSM5599718 GSM5599719 GSM5599720 GSM5599721 GSM5599722 GSM5599723 GSM5599724 GSM5599725 mouse polyA selected RNA samples GSM5599726 GSM5599727 GSM5599728 GSM5599729 GSM5599730 GSM5599731 GSM5599732 GSM5599733 and mouse MII oocyte Input sample GSM5599734. Trimmed reads were aligned to the reference genome mm10 for mouse danRer11 for zebrafish using HISAT2 v2.1.0 with the parameter \u201c 5 8 no mixed no discordant\u201d. Multiply aligned reads were discarded as indicated by HISAT2. PCR duplicates were removed using Samtools v1.9. The read pairs that overlapped with ribosomal RNAs were removed using BEDTools v2.28.0. Genome coverage bigWig files bin size = 10 bp normalized by RPKM were generated by deepTools v3.2.0 bamCoverage with the parameter \u201c bs 10 normalizeUsing RPKM\u201d. Genome build: mm10 for mouse danRer11 for zebrafish Supplementary files format and content: The bigWig files are generated by deepTools v3.2.0 bamCoverage with the bin size=10bp and normalized by RPKM.", "ZF zygote IP 1cell", "For mouse GV oocyte collection 8 wpf C57BL6/N females were injected with 5 units U of pregnant mare serum gonadotropin PMSG. post 48 hours ovaries were dissected and oocytes were isolated by puncturing the follicles. The procedure was carried out in M2 medium Sigma supplemented with 0.2 mM of the cyclic nucleotide phosphodiesterase inhibitor 3 Isobutyl 1 methylxanthine IBMX Sigma to prevent the oocytes from further progress to germinal vesicle breakdown GVBD. The cumulus cells were gently removed by pipetting and the oocytes were briefly exposed to acidic Tyrode's solution Sigma to remove the zona pellucida followed by three washes in M2 medium. For mouse MII oocyte collection 4 wpf 5 wpf C57BL6/N females were injected with 5 U of PMSG followed by 5 U of human chorionic gonadotropin hCG 45 hours later. The oviducts were dissected 20 22 hours later and transferred to a clean dish containing M2 medium. The oviduct ampulla was identified under a stereomicroscope to isolate MII oocytes containing the cumulus mass. The oocytes were treated with 0.3 mg/ml hyaluronidase dissolved in M2 medium to remove the cumulus cells and then exposed to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium. For mouse early embryo collection 8 wpf C57BL/6N females were superovulated by hormone injection 5 U PMSG followed by 5 U hCG 45 hours later and transferred to cages with 8 wpf C57BL/6N males for mating. At 27 28\u2009hours zygote 39 43\u2009hours 2 cell 68 70\u2009hours 8 cell and 92 94\u2009hours blastocyst post hCG administration the female mice were euthanized by cervical dislocation. Embryos were flushed from the reproductive tract into HEPES buffered CZB medium then transferred to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium.", "For zebrafish bulk zygote samples total RNA was extracted from 100 zebrafish zygotes using TRIzol\u2122 Reagent and eluated in 100 \u03bcl of RNase free water. Then 10 \u03bcl and 5 \u03bcl samples were taken for 10 zygote and 5 zygote samples respectively and volumes were adjusted to 12 \u03bcl with nuclease free water. For zebrafish single zygote samples single zygote was manually picked and distributed into 12 \u03bcl of 1X Lysis buffer Takara. For the polyA selected RNA samples of mouse liver and embryonic stem cell WT and Mettl3 KO total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For mouse embryonic stem cell samples the cells were sorted into 12 \u03bcl 1X Lysis buffer using BD FACSMelody\u2122 Cell Sorter BD Biosciences. For mouse oocyte and embryo samples post three washes in M2 medium oocytes and embryos were manually picked and put into 12 \u03bcl 1X Lysis buffer. For picoMeRIP seq experiment NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sheared by sonication incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", null, "developmental stage:Zygote|molecule subtype:rRNA free RNA", "GSM5599725", "GSM5599725: ZF zygote IP 1cell Rep5; Danio rerio; RIP Seq", "GSM5599725", null, "1", "For mouse liver total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For zebrafish zygote mouse MII oocyte and mouse blastocyst samples the single cells were manually picked and sorted into 12 \u03bcl 1X lysis buffer Takara; NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sonicated for 2\u2009\u00d7\u200930\u2009seconds incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", "GEO Accession:GSM5599725", "RIP-Seq", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina NovaSeq 6000", null, "SRP339082", null, "loader:fastq load.py", "ZF_zygote_IP_1cell_Rep5_R1.fastq.gz ZF_zygote_IP_1cell_Rep5_R2.fastq.gz", "fastq fastq", 3660517860.0, 35887430.0, "GSM5599725 r1", "0:51 1:51", "A:858138600;C:924733544;G:1002150083;T:875349961;N:145672", 51, 51, null, null, 858138600, 924733544, 1002150083, 875349961, 145672, "SRX12382357", "SRS10353019", "SRA1301707", "GEO", "Room 2035, Palmer Commons, Department of Computational Medicine and Bioinformatics, University of Michigan", 2, 0.83384, 0.82667, 0.09687, 0.11892, 0.79076, 0.79326, 0.63739, 0.60941, 51, 51, "B", "B", "biological fallback assumption", "illumina", "novaseq_era", "3prime", "poly_a", "nebnext", "bulk", "bulk", "bulk", null, "United States", "2021-09-28", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [66195, "SRR16096232", "SRX12382356", "SRS10353018", "SRP339082", "PRJNA766854", "RNA m6A methylome from zebrafish zygotes mouse liver polyA selected RNAs mouse embryonic stem cells spike in RNA controls and single mouse oocytes and embryos", "GSE184893", "Other", "N6 methyladenosine m6A is a prevalent RNA modification that has key roles in distinct biological processes including meiosis and early embryo development. Despite advances in methodology for m6A mapping the applicability of current methods is limited by the need for large amounts of input material. We develop a sensitive picogram scale m6A RNA immunoprecipitation and sequencing picoMeRIP seq method also suitable for single cell scMeRIP seq. We validate the down scaling of the method by profiling m6A from picogram amounts of mouse liver polyA selected RNA mouse embryonic stem cells and zebrafish zygotes. We provide proof of principle of m6A profiling in single mouse oocytes and preimplantation embryos. Our work opens a new avenue for studying m6A in single cells and scarce cell types in a transcriptome wide manner. Overall design: Transcriptome wide m6A maps were generated from zebrafish and mouse samples as below. 11 zebrafish zygote samples: 3 biological replicates for 10 zygotes 3 biological replicates for 5 zygotes and 5 biological replicates for single zygote. 6 mouse liver polyA selected RNA samples: 10 ng 1 ng and 100 pg; 2 biological replicates for each amount. 9 mouse embryonic stem cell R1 cell line from ATCC SCRC 1011 samples: 1000 cells 100 cells and 10 cells; 3 biological replicates for each amount. 3 spike in RNA control add 0.001 fmol Gaussia luciferase/GLuc with m6A modification and 0.001 fmol Cypridina luciferase/CLuc without xxx modification into the 12 ng polyA selected RNAs related samples: 12 ng polyA selected RNA from mouse liver; 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from wild type/WT mouse embryonic stem cells from Geula S. et al. Science 2015; and 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from Mettl3 knock out/KO mouse embryonic stem cells from Geula S. et al. Science 2015. 12 single mouse oocyte and embryo samples: oocytes at germinal vesicle GV and metaphase II MII stages and embryos at zygote 2 cell 8 cell and blastocyst stages; 2 biological replicates per developmental stage. The pooled Input samples for GV oocyte GSM5746922 zygote GSM5746930 2 cell GSM5746911 and 8 cell GSM5746914 stages can be found in GSE192440.", null, "pubmed:37349523", null, "ZF zygote IP 1cell Rep4", "GSM5599724", null, "tissue:ZF zygote IP 1cell|developmental stage:Zygote|molecule subtype:rRNA free RNA", "ZF zygote IP 1cell Rep4", "Quality of raw reads was assessed using FastQC v0.11.8 and sequencing adapters were trimmed using Cutadapt v1.8.1 if clear adapter signal was detected at the three primeend of the reads. There is no adapter removal for zebrafish zygote samples GSM5599713 GSM5599714 GSM5599715 GSM5599716 GSM5599717 GSM5599718 GSM5599719 GSM5599720 GSM5599721 GSM5599722 GSM5599723 GSM5599724 GSM5599725 mouse polyA selected RNA samples GSM5599726 GSM5599727 GSM5599728 GSM5599729 GSM5599730 GSM5599731 GSM5599732 GSM5599733 and mouse MII oocyte Input sample GSM5599734. Trimmed reads were aligned to the reference genome mm10 for mouse danRer11 for zebrafish using HISAT2 v2.1.0 with the parameter \u201c 5 8 no mixed no discordant\u201d. Multiply aligned reads were discarded as indicated by HISAT2. PCR duplicates were removed using Samtools v1.9. The read pairs that overlapped with ribosomal RNAs were removed using BEDTools v2.28.0. Genome coverage bigWig files bin size = 10 bp normalized by RPKM were generated by deepTools v3.2.0 bamCoverage with the parameter \u201c bs 10 normalizeUsing RPKM\u201d. Genome build: mm10 for mouse danRer11 for zebrafish Supplementary files format and content: The bigWig files are generated by deepTools v3.2.0 bamCoverage with the bin size=10bp and normalized by RPKM.", "ZF zygote IP 1cell", "For mouse GV oocyte collection 8 wpf C57BL6/N females were injected with 5 units U of pregnant mare serum gonadotropin PMSG. post 48 hours ovaries were dissected and oocytes were isolated by puncturing the follicles. The procedure was carried out in M2 medium Sigma supplemented with 0.2 mM of the cyclic nucleotide phosphodiesterase inhibitor 3 Isobutyl 1 methylxanthine IBMX Sigma to prevent the oocytes from further progress to germinal vesicle breakdown GVBD. The cumulus cells were gently removed by pipetting and the oocytes were briefly exposed to acidic Tyrode's solution Sigma to remove the zona pellucida followed by three washes in M2 medium. For mouse MII oocyte collection 4 wpf 5 wpf C57BL6/N females were injected with 5 U of PMSG followed by 5 U of human chorionic gonadotropin hCG 45 hours later. The oviducts were dissected 20 22 hours later and transferred to a clean dish containing M2 medium. The oviduct ampulla was identified under a stereomicroscope to isolate MII oocytes containing the cumulus mass. The oocytes were treated with 0.3 mg/ml hyaluronidase dissolved in M2 medium to remove the cumulus cells and then exposed to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium. For mouse early embryo collection 8 wpf C57BL/6N females were superovulated by hormone injection 5 U PMSG followed by 5 U hCG 45 hours later and transferred to cages with 8 wpf C57BL/6N males for mating. At 27 28\u2009hours zygote 39 43\u2009hours 2 cell 68 70\u2009hours 8 cell and 92 94\u2009hours blastocyst post hCG administration the female mice were euthanized by cervical dislocation. Embryos were flushed from the reproductive tract into HEPES buffered CZB medium then transferred to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium.", "For zebrafish bulk zygote samples total RNA was extracted from 100 zebrafish zygotes using TRIzol\u2122 Reagent and eluated in 100 \u03bcl of RNase free water. Then 10 \u03bcl and 5 \u03bcl samples were taken for 10 zygote and 5 zygote samples respectively and volumes were adjusted to 12 \u03bcl with nuclease free water. For zebrafish single zygote samples single zygote was manually picked and distributed into 12 \u03bcl of 1X Lysis buffer Takara. For the polyA selected RNA samples of mouse liver and embryonic stem cell WT and Mettl3 KO total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For mouse embryonic stem cell samples the cells were sorted into 12 \u03bcl 1X Lysis buffer using BD FACSMelody\u2122 Cell Sorter BD Biosciences. For mouse oocyte and embryo samples post three washes in M2 medium oocytes and embryos were manually picked and put into 12 \u03bcl 1X Lysis buffer. For picoMeRIP seq experiment NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sheared by sonication incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", null, "developmental stage:Zygote|molecule subtype:rRNA free RNA", "GSM5599724", "GSM5599724: ZF zygote IP 1cell Rep4; Danio rerio; RIP Seq", "GSM5599724", null, "1", "For mouse liver total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For zebrafish zygote mouse MII oocyte and mouse blastocyst samples the single cells were manually picked and sorted into 12 \u03bcl 1X lysis buffer Takara; NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sonicated for 2\u2009\u00d7\u200930\u2009seconds incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", "GEO Accession:GSM5599724", "RIP-Seq", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina NovaSeq 6000", null, "SRP339082", null, "loader:fastq load.py", "ZF_zygote_IP_1cell_Rep4_R1.fastq.gz ZF_zygote_IP_1cell_Rep4_R2.fastq.gz", "fastq fastq", 3660205842.0, 35884371.0, "GSM5599724 r1", "0:51 1:51", "A:895738659;C:880838836;G:971118499;T:912364119;N:145729", 51, 51, null, null, 895738659, 880838836, 971118499, 912364119, 145729, "SRX12382356", "SRS10353018", "SRA1301707", "GEO", "Room 2035, Palmer Commons, Department of Computational Medicine and Bioinformatics, University of Michigan", 2, 0.87258, 0.86932, 0.06573, 0.07958, 0.78044, 0.78374, 0.55531, 0.5462, 51, 51, "B", "B", "biological fallback assumption", "illumina", "novaseq_era", "3prime", "poly_a", "nebnext", "bulk", "bulk", "bulk", null, "United States", "2021-09-28", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [66196, "SRR16096231", "SRX12382355", "SRS10353016", "SRP339082", "PRJNA766854", "RNA m6A methylome from zebrafish zygotes mouse liver polyA selected RNAs mouse embryonic stem cells spike in RNA controls and single mouse oocytes and embryos", "GSE184893", "Other", "N6 methyladenosine m6A is a prevalent RNA modification that has key roles in distinct biological processes including meiosis and early embryo development. Despite advances in methodology for m6A mapping the applicability of current methods is limited by the need for large amounts of input material. We develop a sensitive picogram scale m6A RNA immunoprecipitation and sequencing picoMeRIP seq method also suitable for single cell scMeRIP seq. We validate the down scaling of the method by profiling m6A from picogram amounts of mouse liver polyA selected RNA mouse embryonic stem cells and zebrafish zygotes. We provide proof of principle of m6A profiling in single mouse oocytes and preimplantation embryos. Our work opens a new avenue for studying m6A in single cells and scarce cell types in a transcriptome wide manner. Overall design: Transcriptome wide m6A maps were generated from zebrafish and mouse samples as below. 11 zebrafish zygote samples: 3 biological replicates for 10 zygotes 3 biological replicates for 5 zygotes and 5 biological replicates for single zygote. 6 mouse liver polyA selected RNA samples: 10 ng 1 ng and 100 pg; 2 biological replicates for each amount. 9 mouse embryonic stem cell R1 cell line from ATCC SCRC 1011 samples: 1000 cells 100 cells and 10 cells; 3 biological replicates for each amount. 3 spike in RNA control add 0.001 fmol Gaussia luciferase/GLuc with m6A modification and 0.001 fmol Cypridina luciferase/CLuc without xxx modification into the 12 ng polyA selected RNAs related samples: 12 ng polyA selected RNA from mouse liver; 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from wild type/WT mouse embryonic stem cells from Geula S. et al. Science 2015; and 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from Mettl3 knock out/KO mouse embryonic stem cells from Geula S. et al. Science 2015. 12 single mouse oocyte and embryo samples: oocytes at germinal vesicle GV and metaphase II MII stages and embryos at zygote 2 cell 8 cell and blastocyst stages; 2 biological replicates per developmental stage. The pooled Input samples for GV oocyte GSM5746922 zygote GSM5746930 2 cell GSM5746911 and 8 cell GSM5746914 stages can be found in GSE192440.", null, "pubmed:37349523", null, "ZF zygote IP 1cell Rep3", "GSM5599723", null, "tissue:ZF zygote IP 1cell|developmental stage:Zygote|molecule subtype:rRNA free RNA", "ZF zygote IP 1cell Rep3", "Quality of raw reads was assessed using FastQC v0.11.8 and sequencing adapters were trimmed using Cutadapt v1.8.1 if clear adapter signal was detected at the three primeend of the reads. There is no adapter removal for zebrafish zygote samples GSM5599713 GSM5599714 GSM5599715 GSM5599716 GSM5599717 GSM5599718 GSM5599719 GSM5599720 GSM5599721 GSM5599722 GSM5599723 GSM5599724 GSM5599725 mouse polyA selected RNA samples GSM5599726 GSM5599727 GSM5599728 GSM5599729 GSM5599730 GSM5599731 GSM5599732 GSM5599733 and mouse MII oocyte Input sample GSM5599734. Trimmed reads were aligned to the reference genome mm10 for mouse danRer11 for zebrafish using HISAT2 v2.1.0 with the parameter \u201c 5 8 no mixed no discordant\u201d. Multiply aligned reads were discarded as indicated by HISAT2. PCR duplicates were removed using Samtools v1.9. The read pairs that overlapped with ribosomal RNAs were removed using BEDTools v2.28.0. Genome coverage bigWig files bin size = 10 bp normalized by RPKM were generated by deepTools v3.2.0 bamCoverage with the parameter \u201c bs 10 normalizeUsing RPKM\u201d. Genome build: mm10 for mouse danRer11 for zebrafish Supplementary files format and content: The bigWig files are generated by deepTools v3.2.0 bamCoverage with the bin size=10bp and normalized by RPKM.", "ZF zygote IP 1cell", "For mouse GV oocyte collection 8 wpf C57BL6/N females were injected with 5 units U of pregnant mare serum gonadotropin PMSG. post 48 hours ovaries were dissected and oocytes were isolated by puncturing the follicles. The procedure was carried out in M2 medium Sigma supplemented with 0.2 mM of the cyclic nucleotide phosphodiesterase inhibitor 3 Isobutyl 1 methylxanthine IBMX Sigma to prevent the oocytes from further progress to germinal vesicle breakdown GVBD. The cumulus cells were gently removed by pipetting and the oocytes were briefly exposed to acidic Tyrode's solution Sigma to remove the zona pellucida followed by three washes in M2 medium. For mouse MII oocyte collection 4 wpf 5 wpf C57BL6/N females were injected with 5 U of PMSG followed by 5 U of human chorionic gonadotropin hCG 45 hours later. The oviducts were dissected 20 22 hours later and transferred to a clean dish containing M2 medium. The oviduct ampulla was identified under a stereomicroscope to isolate MII oocytes containing the cumulus mass. The oocytes were treated with 0.3 mg/ml hyaluronidase dissolved in M2 medium to remove the cumulus cells and then exposed to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium. For mouse early embryo collection 8 wpf C57BL/6N females were superovulated by hormone injection 5 U PMSG followed by 5 U hCG 45 hours later and transferred to cages with 8 wpf C57BL/6N males for mating. At 27 28\u2009hours zygote 39 43\u2009hours 2 cell 68 70\u2009hours 8 cell and 92 94\u2009hours blastocyst post hCG administration the female mice were euthanized by cervical dislocation. Embryos were flushed from the reproductive tract into HEPES buffered CZB medium then transferred to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium.", "For zebrafish bulk zygote samples total RNA was extracted from 100 zebrafish zygotes using TRIzol\u2122 Reagent and eluated in 100 \u03bcl of RNase free water. Then 10 \u03bcl and 5 \u03bcl samples were taken for 10 zygote and 5 zygote samples respectively and volumes were adjusted to 12 \u03bcl with nuclease free water. For zebrafish single zygote samples single zygote was manually picked and distributed into 12 \u03bcl of 1X Lysis buffer Takara. For the polyA selected RNA samples of mouse liver and embryonic stem cell WT and Mettl3 KO total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For mouse embryonic stem cell samples the cells were sorted into 12 \u03bcl 1X Lysis buffer using BD FACSMelody\u2122 Cell Sorter BD Biosciences. For mouse oocyte and embryo samples post three washes in M2 medium oocytes and embryos were manually picked and put into 12 \u03bcl 1X Lysis buffer. For picoMeRIP seq experiment NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sheared by sonication incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", null, "developmental stage:Zygote|molecule subtype:rRNA free RNA", "GSM5599723", "GSM5599723: ZF zygote IP 1cell Rep3; Danio rerio; RIP Seq", "GSM5599723", null, "1", "For mouse liver total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For zebrafish zygote mouse MII oocyte and mouse blastocyst samples the single cells were manually picked and sorted into 12 \u03bcl 1X lysis buffer Takara; NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sonicated for 2\u2009\u00d7\u200930\u2009seconds incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", "GEO Accession:GSM5599723", "RIP-Seq", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina NovaSeq 6000", null, "SRP339082", null, "loader:fastq load.py", "ZF_zygote_IP_1cell_Rep3_R1.fastq.gz ZF_zygote_IP_1cell_Rep3_R2.fastq.gz", "fastq fastq", 2732409048.0, 26788324.0, "GSM5599723 r1", "0:51 1:51", "A:669345195;C:656715671;G:724415204;T:681825149;N:107829", 51, 51, null, null, 669345195, 656715671, 724415204, 681825149, 107829, "SRX12382355", "SRS10353016", "SRA1301707", "GEO", "Room 2035, Palmer Commons, Department of Computational Medicine and Bioinformatics, University of Michigan", 2, 0.85204, 0.83607, 0.06526, 0.08046, 0.78535, 0.78748, 0.60467, 0.50165, 51, 51, "B", "B", "biological fallback assumption", "illumina", "novaseq_era", "3prime", "poly_a", "nebnext", "bulk", "bulk", "bulk", null, "United States", "2021-09-28", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [66197, "SRR16096230", "SRX12382354", "SRS10353017", "SRP339082", "PRJNA766854", "RNA m6A methylome from zebrafish zygotes mouse liver polyA selected RNAs mouse embryonic stem cells spike in RNA controls and single mouse oocytes and embryos", "GSE184893", "Other", "N6 methyladenosine m6A is a prevalent RNA modification that has key roles in distinct biological processes including meiosis and early embryo development. Despite advances in methodology for m6A mapping the applicability of current methods is limited by the need for large amounts of input material. We develop a sensitive picogram scale m6A RNA immunoprecipitation and sequencing picoMeRIP seq method also suitable for single cell scMeRIP seq. We validate the down scaling of the method by profiling m6A from picogram amounts of mouse liver polyA selected RNA mouse embryonic stem cells and zebrafish zygotes. We provide proof of principle of m6A profiling in single mouse oocytes and preimplantation embryos. Our work opens a new avenue for studying m6A in single cells and scarce cell types in a transcriptome wide manner. Overall design: Transcriptome wide m6A maps were generated from zebrafish and mouse samples as below. 11 zebrafish zygote samples: 3 biological replicates for 10 zygotes 3 biological replicates for 5 zygotes and 5 biological replicates for single zygote. 6 mouse liver polyA selected RNA samples: 10 ng 1 ng and 100 pg; 2 biological replicates for each amount. 9 mouse embryonic stem cell R1 cell line from ATCC SCRC 1011 samples: 1000 cells 100 cells and 10 cells; 3 biological replicates for each amount. 3 spike in RNA control add 0.001 fmol Gaussia luciferase/GLuc with m6A modification and 0.001 fmol Cypridina luciferase/CLuc without xxx modification into the 12 ng polyA selected RNAs related samples: 12 ng polyA selected RNA from mouse liver; 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from wild type/WT mouse embryonic stem cells from Geula S. et al. Science 2015; and 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from Mettl3 knock out/KO mouse embryonic stem cells from Geula S. et al. Science 2015. 12 single mouse oocyte and embryo samples: oocytes at germinal vesicle GV and metaphase II MII stages and embryos at zygote 2 cell 8 cell and blastocyst stages; 2 biological replicates per developmental stage. The pooled Input samples for GV oocyte GSM5746922 zygote GSM5746930 2 cell GSM5746911 and 8 cell GSM5746914 stages can be found in GSE192440.", null, "pubmed:37349523", null, "ZF zygote IP 1cell Rep2", "GSM5599722", null, "tissue:ZF zygote IP 1cell|developmental stage:Zygote|molecule subtype:rRNA free RNA", "ZF zygote IP 1cell Rep2", "Quality of raw reads was assessed using FastQC v0.11.8 and sequencing adapters were trimmed using Cutadapt v1.8.1 if clear adapter signal was detected at the three primeend of the reads. There is no adapter removal for zebrafish zygote samples GSM5599713 GSM5599714 GSM5599715 GSM5599716 GSM5599717 GSM5599718 GSM5599719 GSM5599720 GSM5599721 GSM5599722 GSM5599723 GSM5599724 GSM5599725 mouse polyA selected RNA samples GSM5599726 GSM5599727 GSM5599728 GSM5599729 GSM5599730 GSM5599731 GSM5599732 GSM5599733 and mouse MII oocyte Input sample GSM5599734. Trimmed reads were aligned to the reference genome mm10 for mouse danRer11 for zebrafish using HISAT2 v2.1.0 with the parameter \u201c 5 8 no mixed no discordant\u201d. Multiply aligned reads were discarded as indicated by HISAT2. PCR duplicates were removed using Samtools v1.9. The read pairs that overlapped with ribosomal RNAs were removed using BEDTools v2.28.0. Genome coverage bigWig files bin size = 10 bp normalized by RPKM were generated by deepTools v3.2.0 bamCoverage with the parameter \u201c bs 10 normalizeUsing RPKM\u201d. Genome build: mm10 for mouse danRer11 for zebrafish Supplementary files format and content: The bigWig files are generated by deepTools v3.2.0 bamCoverage with the bin size=10bp and normalized by RPKM.", "ZF zygote IP 1cell", "For mouse GV oocyte collection 8 wpf C57BL6/N females were injected with 5 units U of pregnant mare serum gonadotropin PMSG. post 48 hours ovaries were dissected and oocytes were isolated by puncturing the follicles. The procedure was carried out in M2 medium Sigma supplemented with 0.2 mM of the cyclic nucleotide phosphodiesterase inhibitor 3 Isobutyl 1 methylxanthine IBMX Sigma to prevent the oocytes from further progress to germinal vesicle breakdown GVBD. The cumulus cells were gently removed by pipetting and the oocytes were briefly exposed to acidic Tyrode's solution Sigma to remove the zona pellucida followed by three washes in M2 medium. For mouse MII oocyte collection 4 wpf 5 wpf C57BL6/N females were injected with 5 U of PMSG followed by 5 U of human chorionic gonadotropin hCG 45 hours later. The oviducts were dissected 20 22 hours later and transferred to a clean dish containing M2 medium. The oviduct ampulla was identified under a stereomicroscope to isolate MII oocytes containing the cumulus mass. The oocytes were treated with 0.3 mg/ml hyaluronidase dissolved in M2 medium to remove the cumulus cells and then exposed to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium. For mouse early embryo collection 8 wpf C57BL/6N females were superovulated by hormone injection 5 U PMSG followed by 5 U hCG 45 hours later and transferred to cages with 8 wpf C57BL/6N males for mating. At 27 28\u2009hours zygote 39 43\u2009hours 2 cell 68 70\u2009hours 8 cell and 92 94\u2009hours blastocyst post hCG administration the female mice were euthanized by cervical dislocation. Embryos were flushed from the reproductive tract into HEPES buffered CZB medium then transferred to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium.", "For zebrafish bulk zygote samples total RNA was extracted from 100 zebrafish zygotes using TRIzol\u2122 Reagent and eluated in 100 \u03bcl of RNase free water. Then 10 \u03bcl and 5 \u03bcl samples were taken for 10 zygote and 5 zygote samples respectively and volumes were adjusted to 12 \u03bcl with nuclease free water. For zebrafish single zygote samples single zygote was manually picked and distributed into 12 \u03bcl of 1X Lysis buffer Takara. For the polyA selected RNA samples of mouse liver and embryonic stem cell WT and Mettl3 KO total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For mouse embryonic stem cell samples the cells were sorted into 12 \u03bcl 1X Lysis buffer using BD FACSMelody\u2122 Cell Sorter BD Biosciences. For mouse oocyte and embryo samples post three washes in M2 medium oocytes and embryos were manually picked and put into 12 \u03bcl 1X Lysis buffer. For picoMeRIP seq experiment NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sheared by sonication incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", null, "developmental stage:Zygote|molecule subtype:rRNA free RNA", "GSM5599722", "GSM5599722: ZF zygote IP 1cell Rep2; Danio rerio; RIP Seq", "GSM5599722", null, "1", "For mouse liver total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For zebrafish zygote mouse MII oocyte and mouse blastocyst samples the single cells were manually picked and sorted into 12 \u03bcl 1X lysis buffer Takara; NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sonicated for 2\u2009\u00d7\u200930\u2009seconds incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", "GEO Accession:GSM5599722", "RIP-Seq", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina NovaSeq 6000", null, "SRP339082", null, "loader:fastq load.py", "ZF_zygote_IP_1cell_Rep2_R1.fastq.gz ZF_zygote_IP_1cell_Rep2_R2.fastq.gz", "fastq fastq", 2235668028.0, 21918314.0, "GSM5599722 r1", "0:51 1:51", "A:552522700;C:535947981;G:588168855;T:558939713;N:88779", 51, 51, null, null, 552522700, 535947981, 588168855, 558939713, 88779, "SRX12382354", "SRS10353017", "SRA1301707", "GEO", "Room 2035, Palmer Commons, Department of Computational Medicine and Bioinformatics, University of Michigan", 2, 0.85597, 0.85298, 0.06663, 0.07927, 0.79058, 0.79237, 0.57626, 0.56525, 51, 51, "B", "B", "biological fallback assumption", "illumina", "novaseq_era", "3prime", "poly_a", "nebnext", "bulk", "bulk", "bulk", null, "United States", "2021-09-28", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [66198, "SRR16096229", "SRX12382353", "SRS10353015", "SRP339082", "PRJNA766854", "RNA m6A methylome from zebrafish zygotes mouse liver polyA selected RNAs mouse embryonic stem cells spike in RNA controls and single mouse oocytes and embryos", "GSE184893", "Other", "N6 methyladenosine m6A is a prevalent RNA modification that has key roles in distinct biological processes including meiosis and early embryo development. Despite advances in methodology for m6A mapping the applicability of current methods is limited by the need for large amounts of input material. We develop a sensitive picogram scale m6A RNA immunoprecipitation and sequencing picoMeRIP seq method also suitable for single cell scMeRIP seq. We validate the down scaling of the method by profiling m6A from picogram amounts of mouse liver polyA selected RNA mouse embryonic stem cells and zebrafish zygotes. We provide proof of principle of m6A profiling in single mouse oocytes and preimplantation embryos. Our work opens a new avenue for studying m6A in single cells and scarce cell types in a transcriptome wide manner. Overall design: Transcriptome wide m6A maps were generated from zebrafish and mouse samples as below. 11 zebrafish zygote samples: 3 biological replicates for 10 zygotes 3 biological replicates for 5 zygotes and 5 biological replicates for single zygote. 6 mouse liver polyA selected RNA samples: 10 ng 1 ng and 100 pg; 2 biological replicates for each amount. 9 mouse embryonic stem cell R1 cell line from ATCC SCRC 1011 samples: 1000 cells 100 cells and 10 cells; 3 biological replicates for each amount. 3 spike in RNA control add 0.001 fmol Gaussia luciferase/GLuc with m6A modification and 0.001 fmol Cypridina luciferase/CLuc without xxx modification into the 12 ng polyA selected RNAs related samples: 12 ng polyA selected RNA from mouse liver; 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from wild type/WT mouse embryonic stem cells from Geula S. et al. Science 2015; and 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from Mettl3 knock out/KO mouse embryonic stem cells from Geula S. et al. Science 2015. 12 single mouse oocyte and embryo samples: oocytes at germinal vesicle GV and metaphase II MII stages and embryos at zygote 2 cell 8 cell and blastocyst stages; 2 biological replicates per developmental stage. The pooled Input samples for GV oocyte GSM5746922 zygote GSM5746930 2 cell GSM5746911 and 8 cell GSM5746914 stages can be found in GSE192440.", null, "pubmed:37349523", null, "ZF zygote IP 1cell Rep1", "GSM5599721", null, "tissue:ZF zygote IP 1cell|developmental stage:Zygote|molecule subtype:rRNA free RNA", "ZF zygote IP 1cell Rep1", "Quality of raw reads was assessed using FastQC v0.11.8 and sequencing adapters were trimmed using Cutadapt v1.8.1 if clear adapter signal was detected at the three primeend of the reads. There is no adapter removal for zebrafish zygote samples GSM5599713 GSM5599714 GSM5599715 GSM5599716 GSM5599717 GSM5599718 GSM5599719 GSM5599720 GSM5599721 GSM5599722 GSM5599723 GSM5599724 GSM5599725 mouse polyA selected RNA samples GSM5599726 GSM5599727 GSM5599728 GSM5599729 GSM5599730 GSM5599731 GSM5599732 GSM5599733 and mouse MII oocyte Input sample GSM5599734. Trimmed reads were aligned to the reference genome mm10 for mouse danRer11 for zebrafish using HISAT2 v2.1.0 with the parameter \u201c 5 8 no mixed no discordant\u201d. Multiply aligned reads were discarded as indicated by HISAT2. PCR duplicates were removed using Samtools v1.9. The read pairs that overlapped with ribosomal RNAs were removed using BEDTools v2.28.0. Genome coverage bigWig files bin size = 10 bp normalized by RPKM were generated by deepTools v3.2.0 bamCoverage with the parameter \u201c bs 10 normalizeUsing RPKM\u201d. Genome build: mm10 for mouse danRer11 for zebrafish Supplementary files format and content: The bigWig files are generated by deepTools v3.2.0 bamCoverage with the bin size=10bp and normalized by RPKM.", "ZF zygote IP 1cell", "For mouse GV oocyte collection 8 wpf C57BL6/N females were injected with 5 units U of pregnant mare serum gonadotropin PMSG. post 48 hours ovaries were dissected and oocytes were isolated by puncturing the follicles. The procedure was carried out in M2 medium Sigma supplemented with 0.2 mM of the cyclic nucleotide phosphodiesterase inhibitor 3 Isobutyl 1 methylxanthine IBMX Sigma to prevent the oocytes from further progress to germinal vesicle breakdown GVBD. The cumulus cells were gently removed by pipetting and the oocytes were briefly exposed to acidic Tyrode's solution Sigma to remove the zona pellucida followed by three washes in M2 medium. For mouse MII oocyte collection 4 wpf 5 wpf C57BL6/N females were injected with 5 U of PMSG followed by 5 U of human chorionic gonadotropin hCG 45 hours later. The oviducts were dissected 20 22 hours later and transferred to a clean dish containing M2 medium. The oviduct ampulla was identified under a stereomicroscope to isolate MII oocytes containing the cumulus mass. The oocytes were treated with 0.3 mg/ml hyaluronidase dissolved in M2 medium to remove the cumulus cells and then exposed to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium. For mouse early embryo collection 8 wpf C57BL/6N females were superovulated by hormone injection 5 U PMSG followed by 5 U hCG 45 hours later and transferred to cages with 8 wpf C57BL/6N males for mating. At 27 28\u2009hours zygote 39 43\u2009hours 2 cell 68 70\u2009hours 8 cell and 92 94\u2009hours blastocyst post hCG administration the female mice were euthanized by cervical dislocation. Embryos were flushed from the reproductive tract into HEPES buffered CZB medium then transferred to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium.", "For zebrafish bulk zygote samples total RNA was extracted from 100 zebrafish zygotes using TRIzol\u2122 Reagent and eluated in 100 \u03bcl of RNase free water. Then 10 \u03bcl and 5 \u03bcl samples were taken for 10 zygote and 5 zygote samples respectively and volumes were adjusted to 12 \u03bcl with nuclease free water. For zebrafish single zygote samples single zygote was manually picked and distributed into 12 \u03bcl of 1X Lysis buffer Takara. For the polyA selected RNA samples of mouse liver and embryonic stem cell WT and Mettl3 KO total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For mouse embryonic stem cell samples the cells were sorted into 12 \u03bcl 1X Lysis buffer using BD FACSMelody\u2122 Cell Sorter BD Biosciences. For mouse oocyte and embryo samples post three washes in M2 medium oocytes and embryos were manually picked and put into 12 \u03bcl 1X Lysis buffer. For picoMeRIP seq experiment NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sheared by sonication incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", null, "developmental stage:Zygote|molecule subtype:rRNA free RNA", "GSM5599721", "GSM5599721: ZF zygote IP 1cell Rep1; Danio rerio; RIP Seq", "GSM5599721", null, "1", "For mouse liver total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For zebrafish zygote mouse MII oocyte and mouse blastocyst samples the single cells were manually picked and sorted into 12 \u03bcl 1X lysis buffer Takara; NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sonicated for 2\u2009\u00d7\u200930\u2009seconds incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", "GEO Accession:GSM5599721", "RIP-Seq", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina NovaSeq 6000", null, "SRP339082", null, "loader:fastq load.py", "ZF_zygote_IP_1cell_Rep1_R1.fastq.gz ZF_zygote_IP_1cell_Rep1_R2.fastq.gz", "fastq fastq", 1785761736.0, 17507468.0, "GSM5599721 r1", "0:51 1:51", "A:429081683;C:435813601;G:481376346;T:439419714;N:70392", 51, 51, null, null, 429081683, 435813601, 481376346, 439419714, 70392, "SRX12382353", "SRS10353015", "SRA1301707", "GEO", "Room 2035, Palmer Commons, Department of Computational Medicine and Bioinformatics, University of Michigan", 2, 0.75314, 0.75693, 0.0543, 0.06639, 0.79482, 0.79594, 0.57979, 0.56522, 51, 51, "B", "B", "biological fallback assumption", "illumina", "novaseq_era", "3prime", "poly_a", "nebnext", "bulk", "bulk", "bulk", null, "United States", "2021-09-28", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [66199, "SRR16096228", "SRX12382352", "SRS10353014", "SRP339082", "PRJNA766854", "RNA m6A methylome from zebrafish zygotes mouse liver polyA selected RNAs mouse embryonic stem cells spike in RNA controls and single mouse oocytes and embryos", "GSE184893", "Other", "N6 methyladenosine m6A is a prevalent RNA modification that has key roles in distinct biological processes including meiosis and early embryo development. Despite advances in methodology for m6A mapping the applicability of current methods is limited by the need for large amounts of input material. We develop a sensitive picogram scale m6A RNA immunoprecipitation and sequencing picoMeRIP seq method also suitable for single cell scMeRIP seq. We validate the down scaling of the method by profiling m6A from picogram amounts of mouse liver polyA selected RNA mouse embryonic stem cells and zebrafish zygotes. We provide proof of principle of m6A profiling in single mouse oocytes and preimplantation embryos. Our work opens a new avenue for studying m6A in single cells and scarce cell types in a transcriptome wide manner. Overall design: Transcriptome wide m6A maps were generated from zebrafish and mouse samples as below. 11 zebrafish zygote samples: 3 biological replicates for 10 zygotes 3 biological replicates for 5 zygotes and 5 biological replicates for single zygote. 6 mouse liver polyA selected RNA samples: 10 ng 1 ng and 100 pg; 2 biological replicates for each amount. 9 mouse embryonic stem cell R1 cell line from ATCC SCRC 1011 samples: 1000 cells 100 cells and 10 cells; 3 biological replicates for each amount. 3 spike in RNA control add 0.001 fmol Gaussia luciferase/GLuc with m6A modification and 0.001 fmol Cypridina luciferase/CLuc without xxx modification into the 12 ng polyA selected RNAs related samples: 12 ng polyA selected RNA from mouse liver; 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from wild type/WT mouse embryonic stem cells from Geula S. et al. Science 2015; and 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from Mettl3 knock out/KO mouse embryonic stem cells from Geula S. et al. Science 2015. 12 single mouse oocyte and embryo samples: oocytes at germinal vesicle GV and metaphase II MII stages and embryos at zygote 2 cell 8 cell and blastocyst stages; 2 biological replicates per developmental stage. The pooled Input samples for GV oocyte GSM5746922 zygote GSM5746930 2 cell GSM5746911 and 8 cell GSM5746914 stages can be found in GSE192440.", null, "pubmed:37349523", null, "ZF zygote IP 5cell Rep3", "GSM5599720", null, "tissue:ZF zygote IP 5cell|developmental stage:Zygote|molecule subtype:rRNA free RNA", "ZF zygote IP 5cell Rep3", "Quality of raw reads was assessed using FastQC v0.11.8 and sequencing adapters were trimmed using Cutadapt v1.8.1 if clear adapter signal was detected at the three primeend of the reads. There is no adapter removal for zebrafish zygote samples GSM5599713 GSM5599714 GSM5599715 GSM5599716 GSM5599717 GSM5599718 GSM5599719 GSM5599720 GSM5599721 GSM5599722 GSM5599723 GSM5599724 GSM5599725 mouse polyA selected RNA samples GSM5599726 GSM5599727 GSM5599728 GSM5599729 GSM5599730 GSM5599731 GSM5599732 GSM5599733 and mouse MII oocyte Input sample GSM5599734. Trimmed reads were aligned to the reference genome mm10 for mouse danRer11 for zebrafish using HISAT2 v2.1.0 with the parameter \u201c 5 8 no mixed no discordant\u201d. Multiply aligned reads were discarded as indicated by HISAT2. PCR duplicates were removed using Samtools v1.9. The read pairs that overlapped with ribosomal RNAs were removed using BEDTools v2.28.0. Genome coverage bigWig files bin size = 10 bp normalized by RPKM were generated by deepTools v3.2.0 bamCoverage with the parameter \u201c bs 10 normalizeUsing RPKM\u201d. Genome build: mm10 for mouse danRer11 for zebrafish Supplementary files format and content: The bigWig files are generated by deepTools v3.2.0 bamCoverage with the bin size=10bp and normalized by RPKM.", "ZF zygote IP 5cell", "For mouse GV oocyte collection 8 wpf C57BL6/N females were injected with 5 units U of pregnant mare serum gonadotropin PMSG. post 48 hours ovaries were dissected and oocytes were isolated by puncturing the follicles. The procedure was carried out in M2 medium Sigma supplemented with 0.2 mM of the cyclic nucleotide phosphodiesterase inhibitor 3 Isobutyl 1 methylxanthine IBMX Sigma to prevent the oocytes from further progress to germinal vesicle breakdown GVBD. The cumulus cells were gently removed by pipetting and the oocytes were briefly exposed to acidic Tyrode's solution Sigma to remove the zona pellucida followed by three washes in M2 medium. For mouse MII oocyte collection 4 wpf 5 wpf C57BL6/N females were injected with 5 U of PMSG followed by 5 U of human chorionic gonadotropin hCG 45 hours later. The oviducts were dissected 20 22 hours later and transferred to a clean dish containing M2 medium. The oviduct ampulla was identified under a stereomicroscope to isolate MII oocytes containing the cumulus mass. The oocytes were treated with 0.3 mg/ml hyaluronidase dissolved in M2 medium to remove the cumulus cells and then exposed to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium. For mouse early embryo collection 8 wpf C57BL/6N females were superovulated by hormone injection 5 U PMSG followed by 5 U hCG 45 hours later and transferred to cages with 8 wpf C57BL/6N males for mating. At 27 28\u2009hours zygote 39 43\u2009hours 2 cell 68 70\u2009hours 8 cell and 92 94\u2009hours blastocyst post hCG administration the female mice were euthanized by cervical dislocation. Embryos were flushed from the reproductive tract into HEPES buffered CZB medium then transferred to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium.", "For zebrafish bulk zygote samples total RNA was extracted from 100 zebrafish zygotes using TRIzol\u2122 Reagent and eluated in 100 \u03bcl of RNase free water. Then 10 \u03bcl and 5 \u03bcl samples were taken for 10 zygote and 5 zygote samples respectively and volumes were adjusted to 12 \u03bcl with nuclease free water. For zebrafish single zygote samples single zygote was manually picked and distributed into 12 \u03bcl of 1X Lysis buffer Takara. For the polyA selected RNA samples of mouse liver and embryonic stem cell WT and Mettl3 KO total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For mouse embryonic stem cell samples the cells were sorted into 12 \u03bcl 1X Lysis buffer using BD FACSMelody\u2122 Cell Sorter BD Biosciences. For mouse oocyte and embryo samples post three washes in M2 medium oocytes and embryos were manually picked and put into 12 \u03bcl 1X Lysis buffer. For picoMeRIP seq experiment NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sheared by sonication incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", null, "developmental stage:Zygote|molecule subtype:rRNA free RNA", "GSM5599720", "GSM5599720: ZF zygote IP 5cell Rep3; Danio rerio; RIP Seq", "GSM5599720", null, "1", "For mouse liver total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For zebrafish zygote mouse MII oocyte and mouse blastocyst samples the single cells were manually picked and sorted into 12 \u03bcl 1X lysis buffer Takara; NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sonicated for 2\u2009\u00d7\u200930\u2009seconds incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", "GEO Accession:GSM5599720", "RIP-Seq", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina NovaSeq 6000", null, "SRP339082", null, "loader:fastq load.py", "ZF_zygote_IP_5cell_Rep3_R1.fastq.gz ZF_zygote_IP_5cell_Rep3_R2.fastq.gz", "fastq fastq", 2179659420.0, 21369210.0, "GSM5599720 r1", "0:51 1:51", "A:539092949;C:513857324;G:569620218;T:556940661;N:148268", 51, 51, null, null, 539092949, 513857324, 569620218, 556940661, 148268, "SRX12382352", "SRS10353014", "SRA1301707", "GEO", "Room 2035, Palmer Commons, Department of Computational Medicine and Bioinformatics, University of Michigan", 2, 0.92028, 0.90706, 0.03967, 0.05572, 0.78431, 0.78886, 0.49727, 0.48541, 51, 51, "B", "B", "biological fallback assumption", "illumina", "novaseq_era", "3prime", "poly_a", "nebnext", "bulk", "bulk", "bulk", null, "United States", "2021-09-28", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [66200, "SRR16096227", "SRX12382351", "SRS10353012", "SRP339082", "PRJNA766854", "RNA m6A methylome from zebrafish zygotes mouse liver polyA selected RNAs mouse embryonic stem cells spike in RNA controls and single mouse oocytes and embryos", "GSE184893", "Other", "N6 methyladenosine m6A is a prevalent RNA modification that has key roles in distinct biological processes including meiosis and early embryo development. Despite advances in methodology for m6A mapping the applicability of current methods is limited by the need for large amounts of input material. We develop a sensitive picogram scale m6A RNA immunoprecipitation and sequencing picoMeRIP seq method also suitable for single cell scMeRIP seq. We validate the down scaling of the method by profiling m6A from picogram amounts of mouse liver polyA selected RNA mouse embryonic stem cells and zebrafish zygotes. We provide proof of principle of m6A profiling in single mouse oocytes and preimplantation embryos. Our work opens a new avenue for studying m6A in single cells and scarce cell types in a transcriptome wide manner. Overall design: Transcriptome wide m6A maps were generated from zebrafish and mouse samples as below. 11 zebrafish zygote samples: 3 biological replicates for 10 zygotes 3 biological replicates for 5 zygotes and 5 biological replicates for single zygote. 6 mouse liver polyA selected RNA samples: 10 ng 1 ng and 100 pg; 2 biological replicates for each amount. 9 mouse embryonic stem cell R1 cell line from ATCC SCRC 1011 samples: 1000 cells 100 cells and 10 cells; 3 biological replicates for each amount. 3 spike in RNA control add 0.001 fmol Gaussia luciferase/GLuc with m6A modification and 0.001 fmol Cypridina luciferase/CLuc without xxx modification into the 12 ng polyA selected RNAs related samples: 12 ng polyA selected RNA from mouse liver; 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from wild type/WT mouse embryonic stem cells from Geula S. et al. Science 2015; and 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from Mettl3 knock out/KO mouse embryonic stem cells from Geula S. et al. Science 2015. 12 single mouse oocyte and embryo samples: oocytes at germinal vesicle GV and metaphase II MII stages and embryos at zygote 2 cell 8 cell and blastocyst stages; 2 biological replicates per developmental stage. The pooled Input samples for GV oocyte GSM5746922 zygote GSM5746930 2 cell GSM5746911 and 8 cell GSM5746914 stages can be found in GSE192440.", null, "pubmed:37349523", null, "ZF zygote IP 5cell Rep2", "GSM5599719", null, "tissue:ZF zygote IP 5cell|developmental stage:Zygote|molecule subtype:rRNA free RNA", "ZF zygote IP 5cell Rep2", "Quality of raw reads was assessed using FastQC v0.11.8 and sequencing adapters were trimmed using Cutadapt v1.8.1 if clear adapter signal was detected at the three primeend of the reads. There is no adapter removal for zebrafish zygote samples GSM5599713 GSM5599714 GSM5599715 GSM5599716 GSM5599717 GSM5599718 GSM5599719 GSM5599720 GSM5599721 GSM5599722 GSM5599723 GSM5599724 GSM5599725 mouse polyA selected RNA samples GSM5599726 GSM5599727 GSM5599728 GSM5599729 GSM5599730 GSM5599731 GSM5599732 GSM5599733 and mouse MII oocyte Input sample GSM5599734. Trimmed reads were aligned to the reference genome mm10 for mouse danRer11 for zebrafish using HISAT2 v2.1.0 with the parameter \u201c 5 8 no mixed no discordant\u201d. Multiply aligned reads were discarded as indicated by HISAT2. PCR duplicates were removed using Samtools v1.9. The read pairs that overlapped with ribosomal RNAs were removed using BEDTools v2.28.0. Genome coverage bigWig files bin size = 10 bp normalized by RPKM were generated by deepTools v3.2.0 bamCoverage with the parameter \u201c bs 10 normalizeUsing RPKM\u201d. Genome build: mm10 for mouse danRer11 for zebrafish Supplementary files format and content: The bigWig files are generated by deepTools v3.2.0 bamCoverage with the bin size=10bp and normalized by RPKM.", "ZF zygote IP 5cell", "For mouse GV oocyte collection 8 wpf C57BL6/N females were injected with 5 units U of pregnant mare serum gonadotropin PMSG. post 48 hours ovaries were dissected and oocytes were isolated by puncturing the follicles. The procedure was carried out in M2 medium Sigma supplemented with 0.2 mM of the cyclic nucleotide phosphodiesterase inhibitor 3 Isobutyl 1 methylxanthine IBMX Sigma to prevent the oocytes from further progress to germinal vesicle breakdown GVBD. The cumulus cells were gently removed by pipetting and the oocytes were briefly exposed to acidic Tyrode's solution Sigma to remove the zona pellucida followed by three washes in M2 medium. For mouse MII oocyte collection 4 wpf 5 wpf C57BL6/N females were injected with 5 U of PMSG followed by 5 U of human chorionic gonadotropin hCG 45 hours later. The oviducts were dissected 20 22 hours later and transferred to a clean dish containing M2 medium. The oviduct ampulla was identified under a stereomicroscope to isolate MII oocytes containing the cumulus mass. The oocytes were treated with 0.3 mg/ml hyaluronidase dissolved in M2 medium to remove the cumulus cells and then exposed to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium. For mouse early embryo collection 8 wpf C57BL/6N females were superovulated by hormone injection 5 U PMSG followed by 5 U hCG 45 hours later and transferred to cages with 8 wpf C57BL/6N males for mating. At 27 28\u2009hours zygote 39 43\u2009hours 2 cell 68 70\u2009hours 8 cell and 92 94\u2009hours blastocyst post hCG administration the female mice were euthanized by cervical dislocation. Embryos were flushed from the reproductive tract into HEPES buffered CZB medium then transferred to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium.", "For zebrafish bulk zygote samples total RNA was extracted from 100 zebrafish zygotes using TRIzol\u2122 Reagent and eluated in 100 \u03bcl of RNase free water. Then 10 \u03bcl and 5 \u03bcl samples were taken for 10 zygote and 5 zygote samples respectively and volumes were adjusted to 12 \u03bcl with nuclease free water. For zebrafish single zygote samples single zygote was manually picked and distributed into 12 \u03bcl of 1X Lysis buffer Takara. For the polyA selected RNA samples of mouse liver and embryonic stem cell WT and Mettl3 KO total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For mouse embryonic stem cell samples the cells were sorted into 12 \u03bcl 1X Lysis buffer using BD FACSMelody\u2122 Cell Sorter BD Biosciences. For mouse oocyte and embryo samples post three washes in M2 medium oocytes and embryos were manually picked and put into 12 \u03bcl 1X Lysis buffer. For picoMeRIP seq experiment NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sheared by sonication incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", null, "developmental stage:Zygote|molecule subtype:rRNA free RNA", "GSM5599719", "GSM5599719: ZF zygote IP 5cell Rep2; Danio rerio; RIP Seq", "GSM5599719", null, "1", "For mouse liver total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For zebrafish zygote mouse MII oocyte and mouse blastocyst samples the single cells were manually picked and sorted into 12 \u03bcl 1X lysis buffer Takara; NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sonicated for 2\u2009\u00d7\u200930\u2009seconds incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", "GEO Accession:GSM5599719", "RIP-Seq", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina NovaSeq 6000", null, "SRP339082", null, "loader:fastq load.py", "ZF_zygote_IP_5cell_Rep2_R1.fastq.gz ZF_zygote_IP_5cell_Rep2_R2.fastq.gz", "fastq fastq", 2527184538.0, 24776319.0, "GSM5599719 r1", "0:51 1:51", "A:628307375;C:594702688;G:657847712;T:646151885;N:174878", 51, 51, null, null, 628307375, 594702688, 657847712, 646151885, 174878, "SRX12382351", "SRS10353012", "SRA1301707", "GEO", "Room 2035, Palmer Commons, Department of Computational Medicine and Bioinformatics, University of Michigan", 2, 0.91811, 0.90487, 0.03919, 0.05376, 0.78315, 0.78729, 0.49414, 0.48564, 51, 51, "B", "B", "biological fallback assumption", "illumina", "novaseq_era", "3prime", "poly_a", "nebnext", "bulk", "bulk", "bulk", null, "United States", "2021-09-28", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [66201, "SRR16096226", "SRX12382350", "SRS10353011", "SRP339082", "PRJNA766854", "RNA m6A methylome from zebrafish zygotes mouse liver polyA selected RNAs mouse embryonic stem cells spike in RNA controls and single mouse oocytes and embryos", "GSE184893", "Other", "N6 methyladenosine m6A is a prevalent RNA modification that has key roles in distinct biological processes including meiosis and early embryo development. Despite advances in methodology for m6A mapping the applicability of current methods is limited by the need for large amounts of input material. We develop a sensitive picogram scale m6A RNA immunoprecipitation and sequencing picoMeRIP seq method also suitable for single cell scMeRIP seq. We validate the down scaling of the method by profiling m6A from picogram amounts of mouse liver polyA selected RNA mouse embryonic stem cells and zebrafish zygotes. We provide proof of principle of m6A profiling in single mouse oocytes and preimplantation embryos. Our work opens a new avenue for studying m6A in single cells and scarce cell types in a transcriptome wide manner. Overall design: Transcriptome wide m6A maps were generated from zebrafish and mouse samples as below. 11 zebrafish zygote samples: 3 biological replicates for 10 zygotes 3 biological replicates for 5 zygotes and 5 biological replicates for single zygote. 6 mouse liver polyA selected RNA samples: 10 ng 1 ng and 100 pg; 2 biological replicates for each amount. 9 mouse embryonic stem cell R1 cell line from ATCC SCRC 1011 samples: 1000 cells 100 cells and 10 cells; 3 biological replicates for each amount. 3 spike in RNA control add 0.001 fmol Gaussia luciferase/GLuc with m6A modification and 0.001 fmol Cypridina luciferase/CLuc without xxx modification into the 12 ng polyA selected RNAs related samples: 12 ng polyA selected RNA from mouse liver; 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from wild type/WT mouse embryonic stem cells from Geula S. et al. Science 2015; and 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from Mettl3 knock out/KO mouse embryonic stem cells from Geula S. et al. Science 2015. 12 single mouse oocyte and embryo samples: oocytes at germinal vesicle GV and metaphase II MII stages and embryos at zygote 2 cell 8 cell and blastocyst stages; 2 biological replicates per developmental stage. The pooled Input samples for GV oocyte GSM5746922 zygote GSM5746930 2 cell GSM5746911 and 8 cell GSM5746914 stages can be found in GSE192440.", null, "pubmed:37349523", null, "ZF zygote IP 5cell Rep1", "GSM5599718", null, "tissue:ZF zygote IP 5cell|developmental stage:Zygote|molecule subtype:rRNA free RNA", "ZF zygote IP 5cell Rep1", "Quality of raw reads was assessed using FastQC v0.11.8 and sequencing adapters were trimmed using Cutadapt v1.8.1 if clear adapter signal was detected at the three primeend of the reads. There is no adapter removal for zebrafish zygote samples GSM5599713 GSM5599714 GSM5599715 GSM5599716 GSM5599717 GSM5599718 GSM5599719 GSM5599720 GSM5599721 GSM5599722 GSM5599723 GSM5599724 GSM5599725 mouse polyA selected RNA samples GSM5599726 GSM5599727 GSM5599728 GSM5599729 GSM5599730 GSM5599731 GSM5599732 GSM5599733 and mouse MII oocyte Input sample GSM5599734. Trimmed reads were aligned to the reference genome mm10 for mouse danRer11 for zebrafish using HISAT2 v2.1.0 with the parameter \u201c 5 8 no mixed no discordant\u201d. Multiply aligned reads were discarded as indicated by HISAT2. PCR duplicates were removed using Samtools v1.9. The read pairs that overlapped with ribosomal RNAs were removed using BEDTools v2.28.0. Genome coverage bigWig files bin size = 10 bp normalized by RPKM were generated by deepTools v3.2.0 bamCoverage with the parameter \u201c bs 10 normalizeUsing RPKM\u201d. Genome build: mm10 for mouse danRer11 for zebrafish Supplementary files format and content: The bigWig files are generated by deepTools v3.2.0 bamCoverage with the bin size=10bp and normalized by RPKM.", "ZF zygote IP 5cell", "For mouse GV oocyte collection 8 wpf C57BL6/N females were injected with 5 units U of pregnant mare serum gonadotropin PMSG. post 48 hours ovaries were dissected and oocytes were isolated by puncturing the follicles. The procedure was carried out in M2 medium Sigma supplemented with 0.2 mM of the cyclic nucleotide phosphodiesterase inhibitor 3 Isobutyl 1 methylxanthine IBMX Sigma to prevent the oocytes from further progress to germinal vesicle breakdown GVBD. The cumulus cells were gently removed by pipetting and the oocytes were briefly exposed to acidic Tyrode's solution Sigma to remove the zona pellucida followed by three washes in M2 medium. For mouse MII oocyte collection 4 wpf 5 wpf C57BL6/N females were injected with 5 U of PMSG followed by 5 U of human chorionic gonadotropin hCG 45 hours later. The oviducts were dissected 20 22 hours later and transferred to a clean dish containing M2 medium. The oviduct ampulla was identified under a stereomicroscope to isolate MII oocytes containing the cumulus mass. The oocytes were treated with 0.3 mg/ml hyaluronidase dissolved in M2 medium to remove the cumulus cells and then exposed to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium. For mouse early embryo collection 8 wpf C57BL/6N females were superovulated by hormone injection 5 U PMSG followed by 5 U hCG 45 hours later and transferred to cages with 8 wpf C57BL/6N males for mating. At 27 28\u2009hours zygote 39 43\u2009hours 2 cell 68 70\u2009hours 8 cell and 92 94\u2009hours blastocyst post hCG administration the female mice were euthanized by cervical dislocation. Embryos were flushed from the reproductive tract into HEPES buffered CZB medium then transferred to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium.", "For zebrafish bulk zygote samples total RNA was extracted from 100 zebrafish zygotes using TRIzol\u2122 Reagent and eluated in 100 \u03bcl of RNase free water. Then 10 \u03bcl and 5 \u03bcl samples were taken for 10 zygote and 5 zygote samples respectively and volumes were adjusted to 12 \u03bcl with nuclease free water. For zebrafish single zygote samples single zygote was manually picked and distributed into 12 \u03bcl of 1X Lysis buffer Takara. For the polyA selected RNA samples of mouse liver and embryonic stem cell WT and Mettl3 KO total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For mouse embryonic stem cell samples the cells were sorted into 12 \u03bcl 1X Lysis buffer using BD FACSMelody\u2122 Cell Sorter BD Biosciences. For mouse oocyte and embryo samples post three washes in M2 medium oocytes and embryos were manually picked and put into 12 \u03bcl 1X Lysis buffer. For picoMeRIP seq experiment NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sheared by sonication incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", null, "developmental stage:Zygote|molecule subtype:rRNA free RNA", "GSM5599718", "GSM5599718: ZF zygote IP 5cell Rep1; Danio rerio; RIP Seq", "GSM5599718", null, "1", "For mouse liver total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For zebrafish zygote mouse MII oocyte and mouse blastocyst samples the single cells were manually picked and sorted into 12 \u03bcl 1X lysis buffer Takara; NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sonicated for 2\u2009\u00d7\u200930\u2009seconds incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", "GEO Accession:GSM5599718", "RIP-Seq", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina NovaSeq 6000", null, "SRP339082", null, "loader:fastq load.py", "ZF_zygote_IP_5cell_Rep1_R1.fastq.gz ZF_zygote_IP_5cell_Rep1_R2.fastq.gz", "fastq fastq", 3481342518.0, 34130809.0, "GSM5599718 r1", "0:51 1:51", "A:865680250;C:823420389;G:907146965;T:884854199;N:240715", 51, 51, null, null, 865680250, 823420389, 907146965, 884854199, 240715, "SRX12382350", "SRS10353011", "SRA1301707", "GEO", "Room 2035, Palmer Commons, Department of Computational Medicine and Bioinformatics, University of Michigan", 2, 0.91893, 0.90939, 0.04544, 0.06005, 0.78025, 0.78348, 0.50569, 0.49155, 51, 51, "B", "B", "biological fallback assumption", "illumina", "novaseq_era", "3prime", "poly_a", "nebnext", "bulk", "bulk", "bulk", null, "United States", "2021-09-28", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [66202, "SRR16096225", "SRX12382349", "SRS10353013", "SRP339082", "PRJNA766854", "RNA m6A methylome from zebrafish zygotes mouse liver polyA selected RNAs mouse embryonic stem cells spike in RNA controls and single mouse oocytes and embryos", "GSE184893", "Other", "N6 methyladenosine m6A is a prevalent RNA modification that has key roles in distinct biological processes including meiosis and early embryo development. Despite advances in methodology for m6A mapping the applicability of current methods is limited by the need for large amounts of input material. We develop a sensitive picogram scale m6A RNA immunoprecipitation and sequencing picoMeRIP seq method also suitable for single cell scMeRIP seq. We validate the down scaling of the method by profiling m6A from picogram amounts of mouse liver polyA selected RNA mouse embryonic stem cells and zebrafish zygotes. We provide proof of principle of m6A profiling in single mouse oocytes and preimplantation embryos. Our work opens a new avenue for studying m6A in single cells and scarce cell types in a transcriptome wide manner. Overall design: Transcriptome wide m6A maps were generated from zebrafish and mouse samples as below. 11 zebrafish zygote samples: 3 biological replicates for 10 zygotes 3 biological replicates for 5 zygotes and 5 biological replicates for single zygote. 6 mouse liver polyA selected RNA samples: 10 ng 1 ng and 100 pg; 2 biological replicates for each amount. 9 mouse embryonic stem cell R1 cell line from ATCC SCRC 1011 samples: 1000 cells 100 cells and 10 cells; 3 biological replicates for each amount. 3 spike in RNA control add 0.001 fmol Gaussia luciferase/GLuc with m6A modification and 0.001 fmol Cypridina luciferase/CLuc without xxx modification into the 12 ng polyA selected RNAs related samples: 12 ng polyA selected RNA from mouse liver; 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from wild type/WT mouse embryonic stem cells from Geula S. et al. Science 2015; and 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from Mettl3 knock out/KO mouse embryonic stem cells from Geula S. et al. Science 2015. 12 single mouse oocyte and embryo samples: oocytes at germinal vesicle GV and metaphase II MII stages and embryos at zygote 2 cell 8 cell and blastocyst stages; 2 biological replicates per developmental stage. The pooled Input samples for GV oocyte GSM5746922 zygote GSM5746930 2 cell GSM5746911 and 8 cell GSM5746914 stages can be found in GSE192440.", null, "pubmed:37349523", null, "ZF zygote IP 10cell Rep3", "GSM5599717", null, "tissue:ZF zygote IP 10cell|developmental stage:Zygote|molecule subtype:rRNA free RNA", "ZF zygote IP 10cell Rep3", "Quality of raw reads was assessed using FastQC v0.11.8 and sequencing adapters were trimmed using Cutadapt v1.8.1 if clear adapter signal was detected at the three primeend of the reads. There is no adapter removal for zebrafish zygote samples GSM5599713 GSM5599714 GSM5599715 GSM5599716 GSM5599717 GSM5599718 GSM5599719 GSM5599720 GSM5599721 GSM5599722 GSM5599723 GSM5599724 GSM5599725 mouse polyA selected RNA samples GSM5599726 GSM5599727 GSM5599728 GSM5599729 GSM5599730 GSM5599731 GSM5599732 GSM5599733 and mouse MII oocyte Input sample GSM5599734. Trimmed reads were aligned to the reference genome mm10 for mouse danRer11 for zebrafish using HISAT2 v2.1.0 with the parameter \u201c 5 8 no mixed no discordant\u201d. Multiply aligned reads were discarded as indicated by HISAT2. PCR duplicates were removed using Samtools v1.9. The read pairs that overlapped with ribosomal RNAs were removed using BEDTools v2.28.0. Genome coverage bigWig files bin size = 10 bp normalized by RPKM were generated by deepTools v3.2.0 bamCoverage with the parameter \u201c bs 10 normalizeUsing RPKM\u201d. Genome build: mm10 for mouse danRer11 for zebrafish Supplementary files format and content: The bigWig files are generated by deepTools v3.2.0 bamCoverage with the bin size=10bp and normalized by RPKM.", "ZF zygote IP 10cell", "For mouse GV oocyte collection 8 wpf C57BL6/N females were injected with 5 units U of pregnant mare serum gonadotropin PMSG. post 48 hours ovaries were dissected and oocytes were isolated by puncturing the follicles. The procedure was carried out in M2 medium Sigma supplemented with 0.2 mM of the cyclic nucleotide phosphodiesterase inhibitor 3 Isobutyl 1 methylxanthine IBMX Sigma to prevent the oocytes from further progress to germinal vesicle breakdown GVBD. The cumulus cells were gently removed by pipetting and the oocytes were briefly exposed to acidic Tyrode's solution Sigma to remove the zona pellucida followed by three washes in M2 medium. For mouse MII oocyte collection 4 wpf 5 wpf C57BL6/N females were injected with 5 U of PMSG followed by 5 U of human chorionic gonadotropin hCG 45 hours later. The oviducts were dissected 20 22 hours later and transferred to a clean dish containing M2 medium. The oviduct ampulla was identified under a stereomicroscope to isolate MII oocytes containing the cumulus mass. The oocytes were treated with 0.3 mg/ml hyaluronidase dissolved in M2 medium to remove the cumulus cells and then exposed to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium. For mouse early embryo collection 8 wpf C57BL/6N females were superovulated by hormone injection 5 U PMSG followed by 5 U hCG 45 hours later and transferred to cages with 8 wpf C57BL/6N males for mating. At 27 28\u2009hours zygote 39 43\u2009hours 2 cell 68 70\u2009hours 8 cell and 92 94\u2009hours blastocyst post hCG administration the female mice were euthanized by cervical dislocation. Embryos were flushed from the reproductive tract into HEPES buffered CZB medium then transferred to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium.", "For zebrafish bulk zygote samples total RNA was extracted from 100 zebrafish zygotes using TRIzol\u2122 Reagent and eluated in 100 \u03bcl of RNase free water. Then 10 \u03bcl and 5 \u03bcl samples were taken for 10 zygote and 5 zygote samples respectively and volumes were adjusted to 12 \u03bcl with nuclease free water. For zebrafish single zygote samples single zygote was manually picked and distributed into 12 \u03bcl of 1X Lysis buffer Takara. For the polyA selected RNA samples of mouse liver and embryonic stem cell WT and Mettl3 KO total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For mouse embryonic stem cell samples the cells were sorted into 12 \u03bcl 1X Lysis buffer using BD FACSMelody\u2122 Cell Sorter BD Biosciences. For mouse oocyte and embryo samples post three washes in M2 medium oocytes and embryos were manually picked and put into 12 \u03bcl 1X Lysis buffer. For picoMeRIP seq experiment NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sheared by sonication incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", null, "developmental stage:Zygote|molecule subtype:rRNA free RNA", "GSM5599717", "GSM5599717: ZF zygote IP 10cell Rep3; Danio rerio; RIP Seq", "GSM5599717", null, "1", "For mouse liver total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For zebrafish zygote mouse MII oocyte and mouse blastocyst samples the single cells were manually picked and sorted into 12 \u03bcl 1X lysis buffer Takara; NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sonicated for 2\u2009\u00d7\u200930\u2009seconds incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", "GEO Accession:GSM5599717", "RIP-Seq", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina NovaSeq 6000", null, "SRP339082", null, "loader:fastq load.py", "ZF_zygote_IP_10cell_Rep3_R1.fastq.gz ZF_zygote_IP_10cell_Rep3_R2.fastq.gz", "fastq fastq", 2172362850.0, 21297675.0, "GSM5599717 r1", "0:51 1:51", "A:536043605;C:513512883;G:568368414;T:554284996;N:152952", 51, 51, null, null, 536043605, 513512883, 568368414, 554284996, 152952, "SRX12382349", "SRS10353013", "SRA1301707", "GEO", "Room 2035, Palmer Commons, Department of Computational Medicine and Bioinformatics, University of Michigan", 2, 0.91859, 0.90522, 0.03736, 0.05214, 0.78715, 0.79129, 0.49736, 0.48681, 51, 51, "B", "B", "biological fallback assumption", "illumina", "novaseq_era", "3prime", "poly_a", "nebnext", "bulk", "bulk", "bulk", null, "United States", "2021-09-28", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [66203, "SRR16096224", "SRX12382348", "SRS10353010", "SRP339082", "PRJNA766854", "RNA m6A methylome from zebrafish zygotes mouse liver polyA selected RNAs mouse embryonic stem cells spike in RNA controls and single mouse oocytes and embryos", "GSE184893", "Other", "N6 methyladenosine m6A is a prevalent RNA modification that has key roles in distinct biological processes including meiosis and early embryo development. Despite advances in methodology for m6A mapping the applicability of current methods is limited by the need for large amounts of input material. We develop a sensitive picogram scale m6A RNA immunoprecipitation and sequencing picoMeRIP seq method also suitable for single cell scMeRIP seq. We validate the down scaling of the method by profiling m6A from picogram amounts of mouse liver polyA selected RNA mouse embryonic stem cells and zebrafish zygotes. We provide proof of principle of m6A profiling in single mouse oocytes and preimplantation embryos. Our work opens a new avenue for studying m6A in single cells and scarce cell types in a transcriptome wide manner. Overall design: Transcriptome wide m6A maps were generated from zebrafish and mouse samples as below. 11 zebrafish zygote samples: 3 biological replicates for 10 zygotes 3 biological replicates for 5 zygotes and 5 biological replicates for single zygote. 6 mouse liver polyA selected RNA samples: 10 ng 1 ng and 100 pg; 2 biological replicates for each amount. 9 mouse embryonic stem cell R1 cell line from ATCC SCRC 1011 samples: 1000 cells 100 cells and 10 cells; 3 biological replicates for each amount. 3 spike in RNA control add 0.001 fmol Gaussia luciferase/GLuc with m6A modification and 0.001 fmol Cypridina luciferase/CLuc without xxx modification into the 12 ng polyA selected RNAs related samples: 12 ng polyA selected RNA from mouse liver; 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from wild type/WT mouse embryonic stem cells from Geula S. et al. Science 2015; and 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from Mettl3 knock out/KO mouse embryonic stem cells from Geula S. et al. Science 2015. 12 single mouse oocyte and embryo samples: oocytes at germinal vesicle GV and metaphase II MII stages and embryos at zygote 2 cell 8 cell and blastocyst stages; 2 biological replicates per developmental stage. The pooled Input samples for GV oocyte GSM5746922 zygote GSM5746930 2 cell GSM5746911 and 8 cell GSM5746914 stages can be found in GSE192440.", null, "pubmed:37349523", null, "ZF zygote IP 10cell Rep2", "GSM5599716", null, "tissue:ZF zygote IP 10cell|developmental stage:Zygote|molecule subtype:rRNA free RNA", "ZF zygote IP 10cell Rep2", "Quality of raw reads was assessed using FastQC v0.11.8 and sequencing adapters were trimmed using Cutadapt v1.8.1 if clear adapter signal was detected at the three primeend of the reads. There is no adapter removal for zebrafish zygote samples GSM5599713 GSM5599714 GSM5599715 GSM5599716 GSM5599717 GSM5599718 GSM5599719 GSM5599720 GSM5599721 GSM5599722 GSM5599723 GSM5599724 GSM5599725 mouse polyA selected RNA samples GSM5599726 GSM5599727 GSM5599728 GSM5599729 GSM5599730 GSM5599731 GSM5599732 GSM5599733 and mouse MII oocyte Input sample GSM5599734. Trimmed reads were aligned to the reference genome mm10 for mouse danRer11 for zebrafish using HISAT2 v2.1.0 with the parameter \u201c 5 8 no mixed no discordant\u201d. Multiply aligned reads were discarded as indicated by HISAT2. PCR duplicates were removed using Samtools v1.9. The read pairs that overlapped with ribosomal RNAs were removed using BEDTools v2.28.0. Genome coverage bigWig files bin size = 10 bp normalized by RPKM were generated by deepTools v3.2.0 bamCoverage with the parameter \u201c bs 10 normalizeUsing RPKM\u201d. Genome build: mm10 for mouse danRer11 for zebrafish Supplementary files format and content: The bigWig files are generated by deepTools v3.2.0 bamCoverage with the bin size=10bp and normalized by RPKM.", "ZF zygote IP 10cell", "For mouse GV oocyte collection 8 wpf C57BL6/N females were injected with 5 units U of pregnant mare serum gonadotropin PMSG. post 48 hours ovaries were dissected and oocytes were isolated by puncturing the follicles. The procedure was carried out in M2 medium Sigma supplemented with 0.2 mM of the cyclic nucleotide phosphodiesterase inhibitor 3 Isobutyl 1 methylxanthine IBMX Sigma to prevent the oocytes from further progress to germinal vesicle breakdown GVBD. The cumulus cells were gently removed by pipetting and the oocytes were briefly exposed to acidic Tyrode's solution Sigma to remove the zona pellucida followed by three washes in M2 medium. For mouse MII oocyte collection 4 wpf 5 wpf C57BL6/N females were injected with 5 U of PMSG followed by 5 U of human chorionic gonadotropin hCG 45 hours later. The oviducts were dissected 20 22 hours later and transferred to a clean dish containing M2 medium. The oviduct ampulla was identified under a stereomicroscope to isolate MII oocytes containing the cumulus mass. The oocytes were treated with 0.3 mg/ml hyaluronidase dissolved in M2 medium to remove the cumulus cells and then exposed to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium. For mouse early embryo collection 8 wpf C57BL/6N females were superovulated by hormone injection 5 U PMSG followed by 5 U hCG 45 hours later and transferred to cages with 8 wpf C57BL/6N males for mating. At 27 28\u2009hours zygote 39 43\u2009hours 2 cell 68 70\u2009hours 8 cell and 92 94\u2009hours blastocyst post hCG administration the female mice were euthanized by cervical dislocation. Embryos were flushed from the reproductive tract into HEPES buffered CZB medium then transferred to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium.", "For zebrafish bulk zygote samples total RNA was extracted from 100 zebrafish zygotes using TRIzol\u2122 Reagent and eluated in 100 \u03bcl of RNase free water. Then 10 \u03bcl and 5 \u03bcl samples were taken for 10 zygote and 5 zygote samples respectively and volumes were adjusted to 12 \u03bcl with nuclease free water. For zebrafish single zygote samples single zygote was manually picked and distributed into 12 \u03bcl of 1X Lysis buffer Takara. For the polyA selected RNA samples of mouse liver and embryonic stem cell WT and Mettl3 KO total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For mouse embryonic stem cell samples the cells were sorted into 12 \u03bcl 1X Lysis buffer using BD FACSMelody\u2122 Cell Sorter BD Biosciences. For mouse oocyte and embryo samples post three washes in M2 medium oocytes and embryos were manually picked and put into 12 \u03bcl 1X Lysis buffer. For picoMeRIP seq experiment NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sheared by sonication incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", null, "developmental stage:Zygote|molecule subtype:rRNA free RNA", "GSM5599716", "GSM5599716: ZF zygote IP 10cell Rep2; Danio rerio; RIP Seq", "GSM5599716", null, "1", "For mouse liver total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For zebrafish zygote mouse MII oocyte and mouse blastocyst samples the single cells were manually picked and sorted into 12 \u03bcl 1X lysis buffer Takara; NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sonicated for 2\u2009\u00d7\u200930\u2009seconds incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", "GEO Accession:GSM5599716", "RIP-Seq", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina NovaSeq 6000", null, "SRP339082", null, "loader:fastq load.py", "ZF_zygote_IP_10cell_Rep2_R1.fastq.gz ZF_zygote_IP_10cell_Rep2_R2.fastq.gz", "fastq fastq", 1840839492.0, 18047446.0, "GSM5599716 r1", "0:51 1:51", "A:454385290;C:437082102;G:482351138;T:466893642;N:127320", 51, 51, null, null, 454385290, 437082102, 482351138, 466893642, 127320, "SRX12382348", "SRS10353010", "SRA1301707", "GEO", "Room 2035, Palmer Commons, Department of Computational Medicine and Bioinformatics, University of Michigan", 2, 0.92217, 0.90956, 0.04012, 0.05483, 0.78819, 0.79318, 0.49756, 0.48256, 51, 51, "B", "B", "biological fallback assumption", "illumina", "novaseq_era", "3prime", "poly_a", "nebnext", "bulk", "bulk", "bulk", null, "United States", "2021-09-28", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [66204, "SRR16096223", "SRX12382347", "SRS10353009", "SRP339082", "PRJNA766854", "RNA m6A methylome from zebrafish zygotes mouse liver polyA selected RNAs mouse embryonic stem cells spike in RNA controls and single mouse oocytes and embryos", "GSE184893", "Other", "N6 methyladenosine m6A is a prevalent RNA modification that has key roles in distinct biological processes including meiosis and early embryo development. Despite advances in methodology for m6A mapping the applicability of current methods is limited by the need for large amounts of input material. We develop a sensitive picogram scale m6A RNA immunoprecipitation and sequencing picoMeRIP seq method also suitable for single cell scMeRIP seq. We validate the down scaling of the method by profiling m6A from picogram amounts of mouse liver polyA selected RNA mouse embryonic stem cells and zebrafish zygotes. We provide proof of principle of m6A profiling in single mouse oocytes and preimplantation embryos. Our work opens a new avenue for studying m6A in single cells and scarce cell types in a transcriptome wide manner. Overall design: Transcriptome wide m6A maps were generated from zebrafish and mouse samples as below. 11 zebrafish zygote samples: 3 biological replicates for 10 zygotes 3 biological replicates for 5 zygotes and 5 biological replicates for single zygote. 6 mouse liver polyA selected RNA samples: 10 ng 1 ng and 100 pg; 2 biological replicates for each amount. 9 mouse embryonic stem cell R1 cell line from ATCC SCRC 1011 samples: 1000 cells 100 cells and 10 cells; 3 biological replicates for each amount. 3 spike in RNA control add 0.001 fmol Gaussia luciferase/GLuc with m6A modification and 0.001 fmol Cypridina luciferase/CLuc without xxx modification into the 12 ng polyA selected RNAs related samples: 12 ng polyA selected RNA from mouse liver; 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from wild type/WT mouse embryonic stem cells from Geula S. et al. Science 2015; and 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from Mettl3 knock out/KO mouse embryonic stem cells from Geula S. et al. Science 2015. 12 single mouse oocyte and embryo samples: oocytes at germinal vesicle GV and metaphase II MII stages and embryos at zygote 2 cell 8 cell and blastocyst stages; 2 biological replicates per developmental stage. The pooled Input samples for GV oocyte GSM5746922 zygote GSM5746930 2 cell GSM5746911 and 8 cell GSM5746914 stages can be found in GSE192440.", null, "pubmed:37349523", null, "ZF zygote IP 10cell Rep1", "GSM5599715", null, "tissue:ZF zygote IP 10cell|developmental stage:Zygote|molecule subtype:rRNA free RNA", "ZF zygote IP 10cell Rep1", "Quality of raw reads was assessed using FastQC v0.11.8 and sequencing adapters were trimmed using Cutadapt v1.8.1 if clear adapter signal was detected at the three primeend of the reads. There is no adapter removal for zebrafish zygote samples GSM5599713 GSM5599714 GSM5599715 GSM5599716 GSM5599717 GSM5599718 GSM5599719 GSM5599720 GSM5599721 GSM5599722 GSM5599723 GSM5599724 GSM5599725 mouse polyA selected RNA samples GSM5599726 GSM5599727 GSM5599728 GSM5599729 GSM5599730 GSM5599731 GSM5599732 GSM5599733 and mouse MII oocyte Input sample GSM5599734. Trimmed reads were aligned to the reference genome mm10 for mouse danRer11 for zebrafish using HISAT2 v2.1.0 with the parameter \u201c 5 8 no mixed no discordant\u201d. Multiply aligned reads were discarded as indicated by HISAT2. PCR duplicates were removed using Samtools v1.9. The read pairs that overlapped with ribosomal RNAs were removed using BEDTools v2.28.0. Genome coverage bigWig files bin size = 10 bp normalized by RPKM were generated by deepTools v3.2.0 bamCoverage with the parameter \u201c bs 10 normalizeUsing RPKM\u201d. Genome build: mm10 for mouse danRer11 for zebrafish Supplementary files format and content: The bigWig files are generated by deepTools v3.2.0 bamCoverage with the bin size=10bp and normalized by RPKM.", "ZF zygote IP 10cell", "For mouse GV oocyte collection 8 wpf C57BL6/N females were injected with 5 units U of pregnant mare serum gonadotropin PMSG. post 48 hours ovaries were dissected and oocytes were isolated by puncturing the follicles. The procedure was carried out in M2 medium Sigma supplemented with 0.2 mM of the cyclic nucleotide phosphodiesterase inhibitor 3 Isobutyl 1 methylxanthine IBMX Sigma to prevent the oocytes from further progress to germinal vesicle breakdown GVBD. The cumulus cells were gently removed by pipetting and the oocytes were briefly exposed to acidic Tyrode's solution Sigma to remove the zona pellucida followed by three washes in M2 medium. For mouse MII oocyte collection 4 wpf 5 wpf C57BL6/N females were injected with 5 U of PMSG followed by 5 U of human chorionic gonadotropin hCG 45 hours later. The oviducts were dissected 20 22 hours later and transferred to a clean dish containing M2 medium. The oviduct ampulla was identified under a stereomicroscope to isolate MII oocytes containing the cumulus mass. The oocytes were treated with 0.3 mg/ml hyaluronidase dissolved in M2 medium to remove the cumulus cells and then exposed to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium. For mouse early embryo collection 8 wpf C57BL/6N females were superovulated by hormone injection 5 U PMSG followed by 5 U hCG 45 hours later and transferred to cages with 8 wpf C57BL/6N males for mating. At 27 28\u2009hours zygote 39 43\u2009hours 2 cell 68 70\u2009hours 8 cell and 92 94\u2009hours blastocyst post hCG administration the female mice were euthanized by cervical dislocation. Embryos were flushed from the reproductive tract into HEPES buffered CZB medium then transferred to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium.", "For zebrafish bulk zygote samples total RNA was extracted from 100 zebrafish zygotes using TRIzol\u2122 Reagent and eluated in 100 \u03bcl of RNase free water. Then 10 \u03bcl and 5 \u03bcl samples were taken for 10 zygote and 5 zygote samples respectively and volumes were adjusted to 12 \u03bcl with nuclease free water. For zebrafish single zygote samples single zygote was manually picked and distributed into 12 \u03bcl of 1X Lysis buffer Takara. For the polyA selected RNA samples of mouse liver and embryonic stem cell WT and Mettl3 KO total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For mouse embryonic stem cell samples the cells were sorted into 12 \u03bcl 1X Lysis buffer using BD FACSMelody\u2122 Cell Sorter BD Biosciences. For mouse oocyte and embryo samples post three washes in M2 medium oocytes and embryos were manually picked and put into 12 \u03bcl 1X Lysis buffer. For picoMeRIP seq experiment NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sheared by sonication incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", null, "developmental stage:Zygote|molecule subtype:rRNA free RNA", "GSM5599715", "GSM5599715: ZF zygote IP 10cell Rep1; Danio rerio; RIP Seq", "GSM5599715", null, "1", "For mouse liver total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For zebrafish zygote mouse MII oocyte and mouse blastocyst samples the single cells were manually picked and sorted into 12 \u03bcl 1X lysis buffer Takara; NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sonicated for 2\u2009\u00d7\u200930\u2009seconds incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", "GEO Accession:GSM5599715", "RIP-Seq", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina NovaSeq 6000", null, "SRP339082", null, "loader:fastq load.py", "ZF_zygote_IP_10cell_Rep1_R1.fastq.gz ZF_zygote_IP_10cell_Rep1_R2.fastq.gz", "fastq fastq", 2400553680.0, 23534840.0, "GSM5599715 r1", "0:51 1:51", "A:594878203;C:567573525;G:625488388;T:612448386;N:165178", 51, 51, null, null, 594878203, 567573525, 625488388, 612448386, 165178, "SRX12382347", "SRS10353009", "SRA1301707", "GEO", "Room 2035, Palmer Commons, Department of Computational Medicine and Bioinformatics, University of Michigan", 2, 0.91641, 0.908, 0.03979, 0.05394, 0.78455, 0.7892, 0.48862, 0.47478, 51, 51, "B", "B", "biological fallback assumption", "illumina", "novaseq_era", "3prime", "poly_a", "nebnext", "bulk", "bulk", "bulk", null, "United States", "2021-09-28", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [66205, "SRR16096222", "SRX12382346", "SRS10353008", "SRP339082", "PRJNA766854", "RNA m6A methylome from zebrafish zygotes mouse liver polyA selected RNAs mouse embryonic stem cells spike in RNA controls and single mouse oocytes and embryos", "GSE184893", "Other", "N6 methyladenosine m6A is a prevalent RNA modification that has key roles in distinct biological processes including meiosis and early embryo development. Despite advances in methodology for m6A mapping the applicability of current methods is limited by the need for large amounts of input material. We develop a sensitive picogram scale m6A RNA immunoprecipitation and sequencing picoMeRIP seq method also suitable for single cell scMeRIP seq. We validate the down scaling of the method by profiling m6A from picogram amounts of mouse liver polyA selected RNA mouse embryonic stem cells and zebrafish zygotes. We provide proof of principle of m6A profiling in single mouse oocytes and preimplantation embryos. Our work opens a new avenue for studying m6A in single cells and scarce cell types in a transcriptome wide manner. Overall design: Transcriptome wide m6A maps were generated from zebrafish and mouse samples as below. 11 zebrafish zygote samples: 3 biological replicates for 10 zygotes 3 biological replicates for 5 zygotes and 5 biological replicates for single zygote. 6 mouse liver polyA selected RNA samples: 10 ng 1 ng and 100 pg; 2 biological replicates for each amount. 9 mouse embryonic stem cell R1 cell line from ATCC SCRC 1011 samples: 1000 cells 100 cells and 10 cells; 3 biological replicates for each amount. 3 spike in RNA control add 0.001 fmol Gaussia luciferase/GLuc with m6A modification and 0.001 fmol Cypridina luciferase/CLuc without xxx modification into the 12 ng polyA selected RNAs related samples: 12 ng polyA selected RNA from mouse liver; 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from wild type/WT mouse embryonic stem cells from Geula S. et al. Science 2015; and 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from Mettl3 knock out/KO mouse embryonic stem cells from Geula S. et al. Science 2015. 12 single mouse oocyte and embryo samples: oocytes at germinal vesicle GV and metaphase II MII stages and embryos at zygote 2 cell 8 cell and blastocyst stages; 2 biological replicates per developmental stage. The pooled Input samples for GV oocyte GSM5746922 zygote GSM5746930 2 cell GSM5746911 and 8 cell GSM5746914 stages can be found in GSE192440.", null, "pubmed:37349523", null, "ZF zygote Input 10cell Rep2", "GSM5599714", null, "tissue:ZF zygote Input 10cell|developmental stage:Zygote|molecule subtype:rRNA free RNA", "ZF zygote Input 10cell Rep2", "Quality of raw reads was assessed using FastQC v0.11.8 and sequencing adapters were trimmed using Cutadapt v1.8.1 if clear adapter signal was detected at the three primeend of the reads. There is no adapter removal for zebrafish zygote samples GSM5599713 GSM5599714 GSM5599715 GSM5599716 GSM5599717 GSM5599718 GSM5599719 GSM5599720 GSM5599721 GSM5599722 GSM5599723 GSM5599724 GSM5599725 mouse polyA selected RNA samples GSM5599726 GSM5599727 GSM5599728 GSM5599729 GSM5599730 GSM5599731 GSM5599732 GSM5599733 and mouse MII oocyte Input sample GSM5599734. Trimmed reads were aligned to the reference genome mm10 for mouse danRer11 for zebrafish using HISAT2 v2.1.0 with the parameter \u201c 5 8 no mixed no discordant\u201d. Multiply aligned reads were discarded as indicated by HISAT2. PCR duplicates were removed using Samtools v1.9. The read pairs that overlapped with ribosomal RNAs were removed using BEDTools v2.28.0. Genome coverage bigWig files bin size = 10 bp normalized by RPKM were generated by deepTools v3.2.0 bamCoverage with the parameter \u201c bs 10 normalizeUsing RPKM\u201d. Genome build: mm10 for mouse danRer11 for zebrafish Supplementary files format and content: The bigWig files are generated by deepTools v3.2.0 bamCoverage with the bin size=10bp and normalized by RPKM.", "ZF zygote Input 10cell", "For mouse GV oocyte collection 8 wpf C57BL6/N females were injected with 5 units U of pregnant mare serum gonadotropin PMSG. post 48 hours ovaries were dissected and oocytes were isolated by puncturing the follicles. The procedure was carried out in M2 medium Sigma supplemented with 0.2 mM of the cyclic nucleotide phosphodiesterase inhibitor 3 Isobutyl 1 methylxanthine IBMX Sigma to prevent the oocytes from further progress to germinal vesicle breakdown GVBD. The cumulus cells were gently removed by pipetting and the oocytes were briefly exposed to acidic Tyrode's solution Sigma to remove the zona pellucida followed by three washes in M2 medium. For mouse MII oocyte collection 4 wpf 5 wpf C57BL6/N females were injected with 5 U of PMSG followed by 5 U of human chorionic gonadotropin hCG 45 hours later. The oviducts were dissected 20 22 hours later and transferred to a clean dish containing M2 medium. The oviduct ampulla was identified under a stereomicroscope to isolate MII oocytes containing the cumulus mass. The oocytes were treated with 0.3 mg/ml hyaluronidase dissolved in M2 medium to remove the cumulus cells and then exposed to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium. For mouse early embryo collection 8 wpf C57BL/6N females were superovulated by hormone injection 5 U PMSG followed by 5 U hCG 45 hours later and transferred to cages with 8 wpf C57BL/6N males for mating. At 27 28\u2009hours zygote 39 43\u2009hours 2 cell 68 70\u2009hours 8 cell and 92 94\u2009hours blastocyst post hCG administration the female mice were euthanized by cervical dislocation. Embryos were flushed from the reproductive tract into HEPES buffered CZB medium then transferred to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium.", "For zebrafish bulk zygote samples total RNA was extracted from 100 zebrafish zygotes using TRIzol\u2122 Reagent and eluated in 100 \u03bcl of RNase free water. Then 10 \u03bcl and 5 \u03bcl samples were taken for 10 zygote and 5 zygote samples respectively and volumes were adjusted to 12 \u03bcl with nuclease free water. For zebrafish single zygote samples single zygote was manually picked and distributed into 12 \u03bcl of 1X Lysis buffer Takara. For the polyA selected RNA samples of mouse liver and embryonic stem cell WT and Mettl3 KO total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For mouse embryonic stem cell samples the cells were sorted into 12 \u03bcl 1X Lysis buffer using BD FACSMelody\u2122 Cell Sorter BD Biosciences. For mouse oocyte and embryo samples post three washes in M2 medium oocytes and embryos were manually picked and put into 12 \u03bcl 1X Lysis buffer. For picoMeRIP seq experiment NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sheared by sonication incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", null, "developmental stage:Zygote|molecule subtype:rRNA free RNA", "GSM5599714", "GSM5599714: ZF zygote Input 10cell Rep2; Danio rerio; RIP Seq", "GSM5599714", null, "1", "For mouse liver total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For zebrafish zygote mouse MII oocyte and mouse blastocyst samples the single cells were manually picked and sorted into 12 \u03bcl 1X lysis buffer Takara; NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sonicated for 2\u2009\u00d7\u200930\u2009seconds incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", "GEO Accession:GSM5599714", "RIP-Seq", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina NovaSeq 6000", null, "SRP339082", null, "loader:fastq load.py", "ZF_zygote_Input_10cell_Rep2_R1.fastq.gz ZF_zygote_Input_10cell_Rep2_R2.fastq.gz", "fastq fastq", 3842772582.0, 37674241.0, "GSM5599714 r1", "0:51 1:51", "A:843564828;C:1054412690;G:1130769812;T:813758353;N:266899", 51, 51, null, null, 843564828, 1054412690, 1130769812, 813758353, 266899, "SRX12382346", "SRS10353008", "SRA1301707", "GEO", "Room 2035, Palmer Commons, Department of Computational Medicine and Bioinformatics, University of Michigan", 2, 0.91511, 0.90868, 0.32791, 0.38048, 0.89457, 0.89441, 0.81532, 0.75274, 51, 51, "B", "B", "biological fallback assumption", "illumina", "novaseq_era", "3prime", "poly_a", "nebnext", "bulk", "bulk", "bulk", null, "United States", "2021-09-28", "Zygote", "Embryo", "Oocyte", "Reproductive System"], [66206, "SRR16096221", "SRX12382345", "SRS10353007", "SRP339082", "PRJNA766854", "RNA m6A methylome from zebrafish zygotes mouse liver polyA selected RNAs mouse embryonic stem cells spike in RNA controls and single mouse oocytes and embryos", "GSE184893", "Other", "N6 methyladenosine m6A is a prevalent RNA modification that has key roles in distinct biological processes including meiosis and early embryo development. Despite advances in methodology for m6A mapping the applicability of current methods is limited by the need for large amounts of input material. We develop a sensitive picogram scale m6A RNA immunoprecipitation and sequencing picoMeRIP seq method also suitable for single cell scMeRIP seq. We validate the down scaling of the method by profiling m6A from picogram amounts of mouse liver polyA selected RNA mouse embryonic stem cells and zebrafish zygotes. We provide proof of principle of m6A profiling in single mouse oocytes and preimplantation embryos. Our work opens a new avenue for studying m6A in single cells and scarce cell types in a transcriptome wide manner. Overall design: Transcriptome wide m6A maps were generated from zebrafish and mouse samples as below. 11 zebrafish zygote samples: 3 biological replicates for 10 zygotes 3 biological replicates for 5 zygotes and 5 biological replicates for single zygote. 6 mouse liver polyA selected RNA samples: 10 ng 1 ng and 100 pg; 2 biological replicates for each amount. 9 mouse embryonic stem cell R1 cell line from ATCC SCRC 1011 samples: 1000 cells 100 cells and 10 cells; 3 biological replicates for each amount. 3 spike in RNA control add 0.001 fmol Gaussia luciferase/GLuc with m6A modification and 0.001 fmol Cypridina luciferase/CLuc without xxx modification into the 12 ng polyA selected RNAs related samples: 12 ng polyA selected RNA from mouse liver; 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from wild type/WT mouse embryonic stem cells from Geula S. et al. Science 2015; and 6 ng polyA selected RNA from mouse liver and 6 ng polyA selected RNA from Mettl3 knock out/KO mouse embryonic stem cells from Geula S. et al. Science 2015. 12 single mouse oocyte and embryo samples: oocytes at germinal vesicle GV and metaphase II MII stages and embryos at zygote 2 cell 8 cell and blastocyst stages; 2 biological replicates per developmental stage. The pooled Input samples for GV oocyte GSM5746922 zygote GSM5746930 2 cell GSM5746911 and 8 cell GSM5746914 stages can be found in GSE192440.", null, "pubmed:37349523", null, "ZF zygote Input 10cell Rep1", "GSM5599713", null, "tissue:ZF zygote Input 10cell|developmental stage:Zygote|molecule subtype:rRNA free RNA", "ZF zygote Input 10cell Rep1", "Quality of raw reads was assessed using FastQC v0.11.8 and sequencing adapters were trimmed using Cutadapt v1.8.1 if clear adapter signal was detected at the three primeend of the reads. There is no adapter removal for zebrafish zygote samples GSM5599713 GSM5599714 GSM5599715 GSM5599716 GSM5599717 GSM5599718 GSM5599719 GSM5599720 GSM5599721 GSM5599722 GSM5599723 GSM5599724 GSM5599725 mouse polyA selected RNA samples GSM5599726 GSM5599727 GSM5599728 GSM5599729 GSM5599730 GSM5599731 GSM5599732 GSM5599733 and mouse MII oocyte Input sample GSM5599734. Trimmed reads were aligned to the reference genome mm10 for mouse danRer11 for zebrafish using HISAT2 v2.1.0 with the parameter \u201c 5 8 no mixed no discordant\u201d. Multiply aligned reads were discarded as indicated by HISAT2. PCR duplicates were removed using Samtools v1.9. The read pairs that overlapped with ribosomal RNAs were removed using BEDTools v2.28.0. Genome coverage bigWig files bin size = 10 bp normalized by RPKM were generated by deepTools v3.2.0 bamCoverage with the parameter \u201c bs 10 normalizeUsing RPKM\u201d. Genome build: mm10 for mouse danRer11 for zebrafish Supplementary files format and content: The bigWig files are generated by deepTools v3.2.0 bamCoverage with the bin size=10bp and normalized by RPKM.", "ZF zygote Input 10cell", "For mouse GV oocyte collection 8 wpf C57BL6/N females were injected with 5 units U of pregnant mare serum gonadotropin PMSG. post 48 hours ovaries were dissected and oocytes were isolated by puncturing the follicles. The procedure was carried out in M2 medium Sigma supplemented with 0.2 mM of the cyclic nucleotide phosphodiesterase inhibitor 3 Isobutyl 1 methylxanthine IBMX Sigma to prevent the oocytes from further progress to germinal vesicle breakdown GVBD. The cumulus cells were gently removed by pipetting and the oocytes were briefly exposed to acidic Tyrode's solution Sigma to remove the zona pellucida followed by three washes in M2 medium. For mouse MII oocyte collection 4 wpf 5 wpf C57BL6/N females were injected with 5 U of PMSG followed by 5 U of human chorionic gonadotropin hCG 45 hours later. The oviducts were dissected 20 22 hours later and transferred to a clean dish containing M2 medium. The oviduct ampulla was identified under a stereomicroscope to isolate MII oocytes containing the cumulus mass. The oocytes were treated with 0.3 mg/ml hyaluronidase dissolved in M2 medium to remove the cumulus cells and then exposed to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium. For mouse early embryo collection 8 wpf C57BL/6N females were superovulated by hormone injection 5 U PMSG followed by 5 U hCG 45 hours later and transferred to cages with 8 wpf C57BL/6N males for mating. At 27 28\u2009hours zygote 39 43\u2009hours 2 cell 68 70\u2009hours 8 cell and 92 94\u2009hours blastocyst post hCG administration the female mice were euthanized by cervical dislocation. Embryos were flushed from the reproductive tract into HEPES buffered CZB medium then transferred to acidic Tyrode's solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium.", "For zebrafish bulk zygote samples total RNA was extracted from 100 zebrafish zygotes using TRIzol\u2122 Reagent and eluated in 100 \u03bcl of RNase free water. Then 10 \u03bcl and 5 \u03bcl samples were taken for 10 zygote and 5 zygote samples respectively and volumes were adjusted to 12 \u03bcl with nuclease free water. For zebrafish single zygote samples single zygote was manually picked and distributed into 12 \u03bcl of 1X Lysis buffer Takara. For the polyA selected RNA samples of mouse liver and embryonic stem cell WT and Mettl3 KO total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For mouse embryonic stem cell samples the cells were sorted into 12 \u03bcl 1X Lysis buffer using BD FACSMelody\u2122 Cell Sorter BD Biosciences. For mouse oocyte and embryo samples post three washes in M2 medium oocytes and embryos were manually picked and put into 12 \u03bcl 1X Lysis buffer. For picoMeRIP seq experiment NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sheared by sonication incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", null, "developmental stage:Zygote|molecule subtype:rRNA free RNA", "GSM5599713", "GSM5599713: ZF zygote Input 10cell Rep1; Danio rerio; RIP Seq", "GSM5599713", null, "1", "For mouse liver total RNA was extracted using TRIzol\u2122 Reagent Thermo Fisher Scientific and PolyA+ RNA was selected twice by Dynabeads\u2122 mRNA Purification Kit Thermo Fisher Scientific. For zebrafish zygote mouse MII oocyte and mouse blastocyst samples the single cells were manually picked and sorted into 12 \u03bcl 1X lysis buffer Takara; NEBNext\u00ae rRNA Depletion Kit NEB was used to delete DNA and rRNA; RNA samples were sonicated for 2\u2009\u00d7\u200930\u2009seconds incubated with the anti m6A antibody from Millipore ABE572 and precipitated using ethanol. SMART Seq\u00ae Stranded Kit Takara Cat. # 634442 was used to prepare RNA sequencing library. Libraries were quantified with KAPA Library Quantification Kits Roche and the size distribution was checked using TapeStation D1000 ScreenTape Agilent Technologies.", "GEO Accession:GSM5599713", "RIP-Seq", "TRANSCRIPTOMIC", "other", "PAIRED", "ILLUMINA", "Illumina NovaSeq 6000", null, "SRP339082", null, "loader:fastq load.py", "ZF_zygote_Input_10cell_Rep1_R1.fastq.gz ZF_zygote_Input_10cell_Rep1_R2.fastq.gz", "fastq fastq", 4029368016.0, 39503608.0, "GSM5599713 r1", "0:51 1:51", "A:893562237;C:1096986475;G:1180690796;T:857849967;N:278541", 51, 51, null, null, 893562237, 1096986475, 1180690796, 857849967, 278541, "SRX12382345", "SRS10353007", "SRA1301707", "GEO", "Room 2035, Palmer Commons, Department of Computational Medicine and Bioinformatics, University of Michigan", 2, 0.91491, 0.90807, 0.32799, 0.37836, 0.89292, 0.89282, 0.81006, 0.73354, 51, 51, "B", "B", "biological fallback assumption", "illumina", "novaseq_era", "3prime", "poly_a", "nebnext", "bulk", "bulk", "bulk", null, "United States", "2021-09-28", "Zygote", "Embryo", "Oocyte", "Reproductive System"]], "truncated": false, "filtered_table_rows_count": 51, "expanded_columns": [], "expandable_columns": [], "columns": ["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"], "primary_keys": [], "units": {}, "query": {"sql": "select 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 from run_metadata where \"devstage_curation\" = :p0 and \"experiment.library_selection\" = :p1 order by rowid limit 101", "params": {"p0": "Zygote", "p1": "other"}}, "facet_results": {"experiment.library_strategy": {"name": "experiment.library_strategy", "type": "column", "hideable": false, "toggle_url": "/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other", "results": [{"value": "RIP-Seq", "label": "RIP-Seq", "count": 27, "toggle_url": "http://metadata.rnaquarium.org/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other&experiment.library_strategy=RIP-Seq", "selected": false}, {"value": "OTHER", "label": "OTHER", "count": 24, "toggle_url": "http://metadata.rnaquarium.org/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other&experiment.library_strategy=OTHER", "selected": false}], "truncated": false}, "experiment.library_source": {"name": "experiment.library_source", "type": "column", "hideable": false, "toggle_url": "/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other", "results": [{"value": "TRANSCRIPTOMIC", "label": "TRANSCRIPTOMIC", "count": 51, "toggle_url": "http://metadata.rnaquarium.org/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other&experiment.library_source=TRANSCRIPTOMIC", "selected": false}], "truncated": false}, "experiment.library_selection": {"name": "experiment.library_selection", "type": "column", "hideable": false, "toggle_url": "/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other", "results": [{"value": "other", "label": "other", "count": 51, "toggle_url": "http://metadata.rnaquarium.org/metadata/run_metadata.json?devstage_curation=Zygote", "selected": true}], "truncated": false}, "experiment.library_layout": {"name": "experiment.library_layout", "type": "column", "hideable": false, "toggle_url": "/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other", "results": [{"value": "SINGLE", "label": "SINGLE", "count": 27, "toggle_url": "http://metadata.rnaquarium.org/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other&experiment.library_layout=SINGLE", "selected": false}, {"value": "PAIRED", "label": "PAIRED", "count": 24, "toggle_url": "http://metadata.rnaquarium.org/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other&experiment.library_layout=PAIRED", "selected": false}], "truncated": false}, "experiment.platform": {"name": "experiment.platform", "type": "column", "hideable": false, "toggle_url": "/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other", "results": [{"value": "ILLUMINA", "label": "ILLUMINA", "count": 48, "toggle_url": "http://metadata.rnaquarium.org/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other&experiment.platform=ILLUMINA", "selected": false}, {"value": "ION_TORRENT", "label": "ION_TORRENT", "count": 3, "toggle_url": "http://metadata.rnaquarium.org/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other&experiment.platform=ION_TORRENT", "selected": false}], "truncated": false}, "devstage_curation_coarse": {"name": "devstage_curation_coarse", "type": "column", "hideable": false, "toggle_url": "/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other", "results": [{"value": "Embryo", "label": "Embryo", "count": 51, "toggle_url": "http://metadata.rnaquarium.org/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other&devstage_curation_coarse=Embryo", "selected": false}], "truncated": false}, "devstage_curation": {"name": "devstage_curation", "type": "column", "hideable": false, "toggle_url": "/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other", "results": [{"value": "Zygote", "label": "Zygote", "count": 51, "toggle_url": "http://metadata.rnaquarium.org/metadata/run_metadata.json?experiment.library_selection=other", "selected": true}], "truncated": false}, "tissue_curation_coarse": {"name": "tissue_curation_coarse", "type": "column", "hideable": false, "toggle_url": "/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other", "results": [{"value": "All anatomical structures", "label": "All anatomical structures", "count": 29, "toggle_url": "http://metadata.rnaquarium.org/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other&tissue_curation_coarse=All+anatomical+structures", "selected": false}, {"value": "Reproductive System", "label": "Reproductive System", "count": 22, "toggle_url": "http://metadata.rnaquarium.org/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other&tissue_curation_coarse=Reproductive+System", "selected": false}], "truncated": false}, "tissue_curation": {"name": "tissue_curation", "type": "column", "hideable": false, "toggle_url": "/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other", "results": [{"value": "Embryo Imprecise", "label": "Embryo Imprecise", "count": 22, "toggle_url": "http://metadata.rnaquarium.org/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other&tissue_curation=Embryo+Imprecise", "selected": false}, {"value": "Oocyte", "label": "Oocyte", "count": 22, "toggle_url": "http://metadata.rnaquarium.org/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other&tissue_curation=Oocyte", "selected": false}, {"value": "Whole Organism", "label": "Whole Organism", "count": 7, "toggle_url": "http://metadata.rnaquarium.org/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other&tissue_curation=Whole+Organism", "selected": false}], "truncated": false}, "technology": {"name": "technology", "type": "column", "hideable": false, "toggle_url": "/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other", "results": [{"value": "bulk", "label": "bulk", "count": 17, "toggle_url": "http://metadata.rnaquarium.org/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other&technology=bulk", "selected": false}, {"value": "unknown", "label": "unknown", "count": 15, "toggle_url": "http://metadata.rnaquarium.org/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other&technology=unknown", "selected": false}, {"value": "iclip", "label": "iclip", "count": 13, "toggle_url": "http://metadata.rnaquarium.org/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other&technology=iclip", "selected": false}, {"value": "generic-scrnaseq-only", "label": "generic-scrnaseq-only", "count": 6, "toggle_url": "http://metadata.rnaquarium.org/metadata/run_metadata.json?devstage_curation=Zygote&experiment.library_selection=other&technology=generic-scrnaseq-only", "selected": false}], "truncated": false}}, "suggested_facets": [], "next": null, "next_url": null, "private": false, "allow_execute_sql": true, "query_ms": 106.99567499978002}