|
| Status |
Public on Apr 25, 2014 |
| Title |
BRD2_JQ1 |
| Sample type |
SRA |
| |
|
| Source name |
Prosate cancer cells
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: vertebral metastatic lesion derived prostate cancer cells
cell line: VCaP
chip antibody: BRD2 (Bethyl, A302-583A)
|
| Treatment protocol |
For BRD2/3/4 and ERG ChIP-seq experiments (sample21-33) with BET inhibitors, VCaP cells were treated with DMSO, 500 nM JQ1 or I-BET762 for 12hrs. For AR signaling ChIP-seq experiments (sample1-20), VCaP cells were grown in charcoal-stripped serum containing media for 48hrs. followed by 6hrs. pre-treatment with vehicle or 500nM JQ1 or 10µM MDV3100 or 25µM Bicalutamide and then stimulated with 10nM DHT for 12hrs
|
| Growth protocol |
Cells were grown in DMEM containing 10% FBS
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
ChIP was performed using HighCell ChIP kit following manufacturers protocol (Diagenode). ChIP DNA was isolated (IPure Kit, Diagenode) from samples by incubation with the antibody at 4°C overnight followed by wash and reversal of cross-linking.
The ChIP-seq sample preparation for sequencing was performed according to the manufacturer’s instructions (Illumina). ChIP-enriched DNA samples (1-10 ng) were converted to blunt-ended fragments using T4 DNA polymerase, E.coli DNA polymerase I large fragment (Klenow polymerase) and T4 polynuleotide kinase (New England BioLabs, NEB). A single A-base was added to fragment ends by Klenow fragment (3’ to 5’ exo minus; NEB) followed by ligation of Illumina adaptors (Quick ligase, NEB). The adaptor-modified DNA fragments were enriched by PCR using the Illumina Barcode primers and Phusion DNA polymerase (NEB). PCR products were size selected using 3% NuSieve agarose gels (Lonza) followed by gel extraction using QIAEX II reagents (QIAGEN). Libraries were quantified with the Bioanalyzer 2100 (Agilent) and sequenced on the Illumina HiSeq 2000 Sequencer (100 nucleotide read length).
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 1000 |
| |
|
| Description |
JQ1
|
| Data processing |
Basecalls performed using RTA 1.17.21.3 and CASAVA version 1.8.2
Reads were aligned to the HG19 genome using Bowtie2 with all default settings.
Resulting BAM files were sorted and duplicates were removed using samtools
Coverage profiles were calculated using bedtools genomecov as BedGraph files
BedGraph files wer converted to BigWig using BedGraphToBigWig from Kent Utils
Genome_build: hg19
Supplementary_files_format_and_content: Coverage profiles are normalized to sequencing depth
|
| |
|
| Submission date |
Feb 14, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Marcin Piotr Cieslik |
| E-mail(s) |
[email protected]
|
| Organization name |
University of Michigan
|
| Department |
Pathology
|
| Lab |
MCTP
|
| Street address |
500 S State St
|
| City |
Ann Arbor |
| State/province |
Michigan |
| ZIP/Postal code |
48104 |
| Country |
USA |
| |
|
| Platform ID |
GPL15433 |
| Series (2) |
| GSE55062 |
Therapeutic Targeting of BET Bromodomain Proteins in Castration-Resistant Prostate Cancer (ChIP-Seq) |
| GSE55064 |
Therapeutic Targeting of BET Bromodomain Proteins in Castration-Resistant Prostate Cancer |
|
| Relations |
| BioSample |
SAMN02643389 |
| SRA |
SRX471868 |
