|
| Status |
Public on Sep 28, 2013 |
| Title |
GP5dsiRAD21_HNF4A_Rabbit |
| Sample type |
SRA |
| |
|
| Source name |
GP5d
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: GP5d
cell type: colon adenocarcinoma
hgn: HNF4A
antibody: Santa Cruz Biotechnology, Inc. : sc-8987
control sample: GP5dsiRAD21_IgG_Rabbit
sirna: RAD25
number of peaks, fdr < 5%: 4616
|
| Treatment protocol |
5 μl of HiPerfect (QIAGEN) was mixed with 50 nM of siRNA oligos targeting human RAD21 (Thermo Scientific, cat no M-006832-01) or neutral control (QIAGEN, cat no 1027281) in 100 μl of Opti-MEM (Invitrogen), vortexed vigorously, transferred to a 24-well tissue culture plate and incubated for 10 min at room temperature. 80% confluent GP5d cells were trypsinized, washed with PBS and resuspended in culture medium. Cells (∼1–3 × 104) were added on top of the transfection mixture and cultured for 72 hr before harvesting. Knockdown efficiency was monitored using qPCR and western blotting.
|
| Growth protocol |
GP5d (ECACC cat no. 95090715) colon adenocarcinoma cells were cultured in DMEM supplemented with penicillin/streptomycin and 10% FBS under standard culture conditions.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Cells were cross-linked by 1% formaldehyde, and DNA was sonicated to 200- 400 bp fragments. Antibodies (5 µg) were added and collected using protein G Sepharose (GE). Cross-links were reversed and proteins digested by incubation with proteinase K at 65 °C overnight. DNA was then purified using Qiagen PCR purification kit.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
ChIP-seq experiment on cells treated with siRNA to RAD21
|
| Data processing |
Basecalls performed using Casava 1.4
fastq files were split by barcode
reads were aligned to the human genome (hg18) using BWA
peaks were called with MACS 1.4, using the following settings: --mfold=2,6 –keep-dup=1
Genome_build: hg18
Supplementary_files_format_and_content: text files generated using MACS 1.4
|
| |
|
| Submission date |
Sep 27, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Martin Enge |
| E-mail(s) |
martin.enge@ki.se
|
| Organization name |
Karolinska Institute
|
| Department |
Dep of Oncology-Pathology
|
| Street address |
CCK, Z4
|
| City |
Stockholm |
| ZIP/Postal code |
S-171 76 |
| Country |
Sweden |
| |
|
| Platform ID |
GPL11154 |
| Series (1) |
| GSE51234 |
Transcription factor binding in human cells occurs in dense clusters formed around cohesin anchor sites [GP5d/siRAD21 ChIP-Seq experiments] |
|
| Relations |
| BioSample |
SAMN02364572 |
| SRA |
SRX360584 |
