{"database": "metadata", "table": "run_metadata", "rows": [[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"]], "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": ["rowid"], "primary_key_values": ["66201"], "units": {}, "query_ms": 7.029846994555555}