|
| Status |
Public on Dec 01, 2021 |
| Title |
TAC 3 weeks biological replicate 4 RRBS |
| Sample type |
SRA |
| |
|
| Source name |
Isolated cardiac myocytes
|
| Organism |
Mus musculus |
| Characteristics |
treatment: TAC
time: 3 weeks
|
| Treatment protocol |
DNA was isolated from mouse cardiomyocytes 3 days and 3 weeks after transverse aortic constriction (TAC) and CTCF depletion.
|
| Growth protocol |
-
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
DNA from left ventricular isolated adult cardiomyocytes was isolated using the QIAshredder kit (Qiagen).
Initial digestion into CpG rich regions was done by the Msp1 enzyme (NEB). The 3’ → 5’ Klenow exonuclease was used to repair sticky ends and promote poly-A tail synthesis. To facilitate multiplexing, individual TruSeq Illumina adapters were ligated to samples using the QIAseq Methyl Library Kit (Qiagen). Samples were then filtered to enrich for DNA fragments within the desired size range using magnetic AMPure XP beads (Beckman). Bisulfite conversion was performed using the Zymo EZ DNA Methylation-Lightning kit (Zymo).
|
| |
|
| Library strategy |
Bisulfite-Seq |
| Library source |
genomic |
| Library selection |
Reduced Representation |
| Instrument model |
Illumina HiSeq 3000 |
| |
|
| Data processing |
Single-end reads were demultiplexed into sample-specific *.fastq files using custom scripts and then aligned to the mouse genome using the BSseeker2 v2.0.10.
Briefly, we built an mm10-based reference index—simply 40-400bp MspI restriction fragments digested in silico and consistent with our enriched fragment size—using the bs_seeker2-build.py script.
We then aligned single-end reads to the reference index with bs_seeker2-align.py, using the 40-400bp fragment size range and -r flag to indicate RRBS libraries, and allowing up to 5 mismatches. We selected Bowtie2 as the aligner within BSseeker2 for this step.
Samtools was used to sort the resulting .bam files, and methylation was called using bs_seeker2-call_methylation.py, using a 10 read coverage minimum for any given cytosine, and removing all reads that did not undergo full bisulfite conversion.
From the resulting CGmaps, we retained cytosines that exist within a CpG (cytosine followed by guanine) context. These are deposited herein as processed data.
Genome_build: mm10
Supplementary_files_format_and_content: CGmap files
|
| |
|
| Submission date |
Jul 15, 2020 |
| Last update date |
Dec 01, 2021 |
| Contact name |
Tom Vondriska |
| E-mail(s) |
vondriska.web@gmail.com
|
| Organization name |
UCLA
|
| Street address |
650 Charles E Young Dr, BH553
|
| City |
Los Angeles |
| State/province |
CA |
| ZIP/Postal code |
90095 |
| Country |
USA |
| |
|
| Platform ID |
GPL21493 |
| Series (2) |
| GSE154520 |
Temporal Analyses of Cardiac Chromatin Accessibility, DNA Methylation and Epigenomic Structure Reveal Locus-Specific Regulation [RRBS] |
| GSE154521 |
Temporal Analyses of Cardiac Chromatin Accessibility, DNA Methylation and Epigenomic Structure Reveal Locus-Specific Regulation |
|
| Relations |
| BioSample |
SAMN15545633 |
| SRA |
SRX8740608 |
