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Sample GSM5049377 Query DataSets for GSM5049377
Status Public on Jan 29, 2021
Title 50_F_KO_10_RRBS
Sample type SRA
 
Source name Female_KO_10_soleus muscle
Organism Mus musculus
Characteristics strain background: C57BL/6
Sex: Female
genotype: KO
timepoint: 10
tissue: soleus muscle
Extracted molecule genomic DNA
Extraction protocol A single soleus muscle or ~30 mg of powdered quadriceps was homogenized in 400 µl of RLT buffer (Qiagen) with β-mercaptoethanol using steal beads and subjected to 2 x 30 sec at 30 Hz of disruption using the TissueLyser II (Qiagen). RNA and DNA were simultaneously isolated from 350 µl of lysate using the RNA/DNA/miRNA AllPrep kit (Qiagen) as per instructions. Primary muscle cells were lysed directly in RLT buffer and RNA extracted using the RNeasy kit (Qiagen). RNA was eluted in 45 µl of RNAse free water and DNA in 60 µl of elution buffer (Qiagen).
Reduced Representation Bisulfite Sequencing (RRBS) was performed as described (Laker et al., 2017). Briefly, DNA samples were incubated with MpsI enzyme (NEB) in order to fragment DNA at CCGG positions to enrich for CpG regions. Bisulfite conversion was performed using the EZ DNA methylation Kit (Zymo Research) according to the manufacturer’s instructions. The DNA was then PCR amplified and ligated to TruSeq (Illumina) sequencing adapters. Libraries were subjected to 75-bp single-end sequencing on the NextSeq 550 (Illumina). On average 18.3 million reads/sample were aligned
 
Library strategy Bisulfite-Seq
Library source genomic
Library selection Reduced Representation
Instrument model Illumina NextSeq 500
 
Description This sample was used to estimate global DNA methylation levels and mitochondrial DNA copy numbers, but was excluded from all differential methylation analyses.
Data processing reads were preprocessed with Trim Galore v. 0.4.0, a wrapper for cutadapt v. 1.9.1, with the --rrbs flag set
trimmed reads were aligned to the mm10 genome assembly using Bismark v. 0.18.1 with default parameters
CpG methylation calls were summarized using the bismark2bedgraph script which is part of the Bismark software suite
For each CpG, observations from both strands were added together to form an overall methylation level per CpG, and only CpGs covered by eight or more reads in every sample was retained
Genome_build: Genome_build: mm10
Supplementary_files_format_and_content: Supplementary_files_format_and_content: csv file with CpGs in rows and genomic annotation along with number of methylated and unmethylated observations for each sample in columns.
 
Submission date Jan 28, 2021
Last update date Feb 04, 2021
Contact name Lars Roed Ingerslev
E-mail(s) ingerslev@sund.ku.dk
Organization name Copenhagen University
Department NNF Center for Basic Metabolic Research
Lab Integrative Physiology
Street address Blegdamsvej 3B
City Copenhagen
ZIP/Postal code 2200
Country Denmark
 
Platform ID GPL19057
Series (2)
GSE152348 Ablation of DNA-methyltransferase 3A in skeletal muscle does not affect energy metabolism or exercise capacity [RRBS]
GSE152349 Ablation of DNA-methyltransferase 3A in skeletal muscle does not affect energy metabolism or exercise capacity
Relations
BioSample SAMN17619441
SRA SRX9973042

Supplementary file Size Download File type/resource
GSM5049377_CpG_context_50_F_KO_10_RRBS.cov.gz 8.2 Mb (ftp)(http) COV
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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