|
| Status |
Public on Jul 24, 2013 |
| Title |
prolif_Ubc9_ChIPSeq |
| Sample type |
SRA |
| |
|
| Source name |
prolif_Ubc9_ChIPSeq
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: WI38
cell type: primary fibroblasts
status: Proliferative
genotype/variation: infected with pBABE-puro-empty
chip antibodies: (BD transduction)
|
| Treatment protocol |
Infection by retroviral-mediated gene transfer with either pBABE-puro-H-RasG12V or empty vector were performed as previously described (Bischof et al., 2006)
|
| Growth protocol |
Culturing of human diploid WI38 fibroblasts (ATCC) was performed at physiological oxygen concentration of 3% as previously described (Bischof et al., 2006)
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Lysates were clarified from sonicated nuclei and protein-DNA complexes were isolated with antibody.
Libraries were prepared using ChIP-Seq Sample Prep Kit (#IP-102-1001, Illumina) following the manufacturer's protocol with some modifications. Briefly, 10 ng of ChIP enriched DNA or control DNA were end repaired using T4 DNA polymerase, Klenow DNA polymerase and T4 PNK. A single ‘A’ nucleotide was added to the 3’ ends of the blunt DNA fragments with a Klenow fragment (3' to 5'exo minus). The ends of the DNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR (30 sec at 98°C; [10 sec at 98°C, 30 sec at 65°C, 30 sec at 72°C] x 14 cycles; 5 min at 72°C) and then purified using Agencourt AMPure XP beads (#A63881, Beckman). DNA library size selection was performed by excising the 250-350 bp fragments from a 2% agarose gel followed by purification using a QIAquick Gel Extraction Kit (#28906, Qiagen). Prior to analyses DNA libraries were checked for quality and quantified using a 2100 Bioanalyzer (Agilent). The libraries were loaded in the flowcell at 8pM concentration and clusters were generated using the Cbot and sequenced on the Illumina Genome Analyzer IIx as single-end 36 base reads following Illumina’s instructions.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina Genome Analyzer IIx |
| |
|
| Data processing |
Basecalls performed using CASAVA version 1.7 or 1.8
ChIP-seq reads were aligned to the hg19 genome assembly using the Illumina Genome Analyzer Pipeline or Bowtie aligner.
Peaks were called using the Model-based Analysis of ChIP-Seq (MACS 1.3.7) algorithm (http://liulab.dfci.harvard.edu/MACS/). Results were visualized using the UCSC Genome Browser (http://genome.ucsc.edu). Global clustering analysis and quantitative comparisons were performed using seqMINER [(34) (http://bips.ustrasbg. fr/seqminer/].
Wig files: For all samples, aligned sequences were extended to 200bp downstream (with respect to read strand) and allocated into 25bp bins.
Genome_build: hg19
Supplementary_files_format_and_content: bed (macs peaks)
|
| |
|
| Submission date |
Nov 09, 2012 |
| Last update date |
May 15, 2019 |
| Contact name |
Tao YE |
| Organization name |
IGBMC (CNRS/INSERM/UDS)
|
| Street address |
1 rue Laurent Fries
|
| City |
Illkirch |
| ZIP/Postal code |
67404 |
| Country |
France |
| |
|
| Platform ID |
GPL10999 |
| Series (2) |
| GSE42211 |
Genome wide occupancy of SUMO machinery in proliferative and Ras-induced senescent human primary fibroblasts |
| GSE42213 |
SUMO is an Integral and Instructive Component of Chromatin in Cell Growth and Senescence |
|
| Relations |
| SRA |
SRX204165 |
| BioSample |
SAMN01805518 |
