|
| Status |
Public on Mar 23, 2022 |
| Title |
RRBS_Ocun_fibro_5aza0 |
| Sample type |
SRA |
| |
|
| Source name |
Primary dermal fibroblasts
|
| Organism |
Oryctolagus cuniculus |
| Characteristics |
Sex: Female
genome build: oryCun2
treatment: Control (not treated)
tissue: Primary dermal fibroblasts
|
| Treatment protocol |
Fibroblasts were treated with 1 µM final concentration of 5-AzadC (Sigma A3656) for 72 hours and the medium was renewed every day.
|
| Growth protocol |
Primary dermal fibroblasts were grown in DMEM-GlutaMax supplemented with 10% fetal bovine serum and 50U/mL penicillin-streptomycin in a humidified atmosphere containing 5% CO2 at 37°C.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Genomic DNA was extracted using the AllPrep DNA/RNA Mini Kit (Qiagen).
100 ng of genomic DNA were digested 5 hours by MspI (Thermo Scientific), end-repaired and A-tailed with Klenow fragment exo- (Thermo Scientific), and ligated to methylated adapters with T4 DNA ligase (Thermo Scientific) in Tango 1X buffer. Size selection was performed by gel excision from a 3% agarose 0.5X TBE gel to select fragments ranging from 150 to 400 bp. DNA was purified using the MinElute gel extraction kit (Qiagen) and bisulfite-converted twice with the EpiTect bisulfite kit (Qiagen), following the manufacturer’s instructions. Final libraries were amplified using the Pfu Turbo Cx hotstart DNA polymerase (Agilent) with 12 to 14 PCR cycles. The libraries were purified using Agencourt AmpureXP beads (Beckman-Coulter).
|
| |
|
| Library strategy |
Bisulfite-Seq |
| Library source |
genomic |
| Library selection |
Reduced Representation |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Data processing |
Reads were trimmed with Trim Galore v0.4.4 to remove adapter sequences and low-quality ends with a Phred score below 20. Trim Galore was run in –non_directional and –rrbs mode to remove two additional bases artificially introduced at the MspI restriction sites.
Reads were aligned to the corresponding genome with Bismark v0.18.2 with default parameters.
A maximum of two mismatches and an insertion size for paired-end sequences of between 30 and 400 bp were allowed.
Methylation scores were extracted as the ratio of the number of Cs over the total number of Cs and Ts using the Bismark_methylation_extractor. CGs were filtered to have a minimum sequencing depth of 10X.
genome build: hg38, mm10, oryCun2, canFam3, bosTau8, susScr11, galGal6
processed data files format and content: We provide IGV files containing methylation scores for all CGs with at least 10X sequencing depth.
|
| |
|
| Submission date |
May 27, 2021 |
| Last update date |
Mar 23, 2022 |
| Contact name |
Michael Weber |
| Organization name |
CNRS
|
| Department |
UMR7242 Biotechnology and Cell Signalling
|
| Street address |
300 Bd Sebastien Brant
|
| City |
Illkirch |
| ZIP/Postal code |
67412 |
| Country |
France |
| |
|
| Platform ID |
GPL25392 |
| Series (1) |
| GSE175615 |
Conservation and divergence of DNA methylation patterns and functions in vertebrates |
|
| Relations |
| BioSample |
SAMN19365329 |
| SRA |
SRX11003170 |
