GSM3752614: WGBS_WT; Mus musculus; Bisulfite-Seq - SRA

SRX5795080: GSM3752614: WGBS_WT; Mus musculus; Bisulfite-Seq
1 ILLUMINA (Illumina HiSeq 4000) run: 301.6M spots, 60.3G bases, 27.6Gb downloads

Submitted by: NCBI (GEO)

Study: Genome-wide roles of DNA methyltransferases in mouse embryos

PRJNA541237 • SRP195507 • All experiments • All runs

show Abstracthide Abstract

Mouse embryos acquire global DNA methylation of their genome during implantation. However the exact roles of DNA methyltransferases (DNMTs) in embryognesis have not been studied comprehensively. Here we systematically analyze the consequences of genetic inactivation of Dnmt1, Dnmt3a and Dnmt3b on the methylome and transcriptome of mouse embryos and fibroblasts. Overall design: We mapped DNA methylation by Whole Genome Bisulfite Sequencing (WGBS) in two independent WT, Dnmt1-/- and Dnmt3a-/- Dnmt3b-/- (DKO) E8.5 embryos. We also sequenced RRBS libraries from three Dnmt1-/- (D1KO_Rep1-3) and WT (D1WT_Rep1-3) littermate E8.5 embryos, as well as three Dnmt3a-/- Dnmt3b-/- (DKO_Rep1-3) together with two WT (D3abWT_Rep1-2), three Dnmt3a-/+ (D3aHet_Rep1-3), three Dnmt3a-/+ Dnmt3b-/+ (D3abHet_Rep1-3) littermate E8.5 embryos. To study the role of DNMT3A/B and DNMT1 in maintenance methylation, we generated Dnmt3aL2/L2; Dnmt3bL2/L2; CreERT2 and Dnmt1L2/L2; CreERT2 immortalized mouse embryonic fibroblasts (MEFs). The CreERT2 recombinase is activated by tamoxifen treatment to generate Dnmt3a Dnmt3b double conditional knockout (cDKO) and Dnmt1 conditional knockout (D1cKO) MEFs. We performed three independent tamoxifen induction experiments and profiled DNA methylation by RRBS in cells treated with Tamoxifen (Tam_Rep1-3) or not treated with Tamoxifen (noTam_Rep1-3) at various time points of culture (day 23 and day 69 for cDKO MEFs, day 5 and day 7 for D1cKO MEFs). The transcriptome of Dnmt mutant embryos was analyzed by RNA-seq. We sequenced RNA-seq libraries from three Dnmt1-/- (D1KO_Rep1-3) and three WT (D1WT_Rep1-3) littermate E8.5 embryos, as well as six Dnmt3a-/- Dnmt3b-/- E8.5 embryos (DKO_Rep1-6) together with two WT (D3abWT_Rep1-2) and four Dnmt3a-/+ (D3aHet_Rep1-4) littermate controls.

Sample: WGBS_WT_Rep1

SAMN11580853 • SRS4726455 • All experiments • All runs

Organism: Mus musculus

Library:

Instrument: Illumina HiSeq 4000

Strategy: Bisulfite-Seq

Source: GENOMIC

Selection: RANDOM

Layout: PAIRED

Construction protocol: WGBS: Genomic DNA was extracted with the AllPrep DNA/RNA Mini Kit (Qiagen). WGBS: 50 ng of genomic DNA were fragmented to 350 bp using a Covaris E220 sonicator. DNA was bisulfite-converted with the EZ DNA Methylation-Gold kit (Zymo Research) and WGBS libraries were prepared using the Accel-NGS Methyl-Seq DNA Library Kit (Swift Biosciences) according to the manufacturer's instructions with 7 PCR cycles for the final amplification. The libraries were purified using Ampure XP beads (Beckman Coulter). RRBS: Genomic DNA was extracted with the AllPrep DNA/RNA Mini Kit (Qiagen). RRBS: RRBS libraries were prepared as described in Auclair et al., Genome Biol 15:545 (2014). Briefly, we digested genomic DNA 5 hours with MspI (Thermo Scientific), followed by end-repair and A-tailing (Klenow fragment, Thermo Scientific) and ligation to methylated adapters (T4 DNA ligase, Thermo Scientific) in Tango 1X buffer. Fragments between 150 and 400 bp were excised from a 3% agarose 0.5X TBE gel with the MinElute kit (Qiagen) and bisulfite-converted with the EpiTect Bisulfite kit (Qiagen) using two consecutive rounds of conversion. Final RRBS libraries were amplified with 14-16 cycles of PCR. The libraries were purified using Ampure XP beads (Beckman Coulter). RNA-seq: Total RNA was extracted with the AllPrep DNA/RNA Mini Kit (Qiagen). RNA-seq: RNA-Seq libraries were prepared with the TruSeq Stranded Total RNA Sample Prep Kit (Illumina) with Ribo-Zero depletion according to the manufacturer's instructions.

Experiment attributes:

GEO Accession: GSM3752614

Links:

NCBI link: NCBI Entrez (gds)

Runs: 1 run, 301.6M spots, 60.3G bases, 27.6Gb

Run	# of Spots	# of Bases	Size	Published
SRR9016926	301,591,204	60.3G	27.6Gb	2020-05-21

ID:: 7791543

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