|
| Status |
Public on Aug 28, 2012 |
| Title |
Keratinocytes |
| Sample type |
SRA |
| |
|
| Source name |
Keratinocytes
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Human keratinocytes
laboratory of generation: NA
reprogramming experiment: NA
reprogramming factors: NA
passage number: 3
|
| Treatment protocol |
Cells were treated with mTeSR1 and Activin-A or BMP4 or media only for 5 days then collected with TrypLE (Invitrogen).
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Bisulfite padlock probes capture and sequencing (Diep et al, Nature Methods 2012)
Genomic DNA was extracted using the ALLPrep DNA/RNA Mini kit (Qiagen) and QIAamp DNA Micro kit (Qiagen). The bisulfite conversion and capture reactions were carried out on 1-1.2 g of each sample using previously established protocols (13, 28, 29). Briefly, DNA was bisulfite converted using the EZ-96Methylation Gold Kit (Zymo Research). About 200-300 ng of converted gDNA from each sample was captured using prepared padlock probe oligonucleotides, resulting in a circular DNA library of targeted CpG sites. Each DNA library was amplified using limited cycle PCR and size-selected on polyacrylamide gels before sequencing.
|
| |
|
| Library strategy |
Bisulfite-Seq |
| Library source |
genomic |
| Library selection |
RANDOM |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Data processing |
Illumina OLB 1.9.3 (for HiSeq run) and OLB 1.8.0 (for GA run) were used for base-calling
Adaptor sequences (27bp from 5' end) were trimmed from bisulfite reads prior to mapping
Bisulfite reads were mapped using the protocol from Diep et al, Nature Methods 2012. Briefly, all 'C's were replaced by 'T's in reads and mapped to a reference with all 'C's converted to 'T's. After mapping, the modified reads were replaced by the original reads then pileup files were generated using SAMTOOLS
Methylation fractions were calculated as the number of cytosines over the total thymines and cytosines for CpGs where at least 90% of base called were thymines and cytosines
CpG sites with at least 10 depth of coverage were reported in final processed file (BED format)
Genome_build: hg18
Supplementary_files_format_and_content: BED formated (tab-delimited) text files include chromosome position and methylation fraction value for each CpG in each sample.
|
| |
|
| Submission date |
Aug 24, 2012 |
| Last update date |
May 15, 2019 |
| Contact name |
Kun Zhang |
| E-mail(s) |
k4zhang@ucsd.edu
|
| Organization name |
University of California, San Diego
|
| Department |
Bioengineering
|
| Lab |
Integrative Genomics Laboratory
|
| Street address |
9500 Gilman Dr Mailcode: 0412
|
| City |
La Jolla |
| State/province |
CA |
| ZIP/Postal code |
92093 |
| Country |
USA |
| |
|
| Platform ID |
GPL11154 |
| Series (1) |
| GSE40372 |
Identification of a specific reprogramming-associated epigenetic signature in human induced pluripotent stem cells |
|
| Relations |
| SRA |
SRX180789 |
| BioSample |
SAMN01129776 |
