|
| Status |
Public on Jan 23, 2014 |
| Title |
RING1B_CST_sh3_ChIPSeq |
| Sample type |
SRA |
| |
|
| Source name |
DU145 prostate cancer cells
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: DU145 prostate cancer cells
genotype/variation: EED knockdown
chip antibody: Anti-RING1B [rabbit, Cell Signaling, cat# 5694, lot# 1]
|
| Treatment protocol |
stable cells were selected with puromycin
|
| Growth protocol |
Du145 cells were growth following ATCC instruction
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Lysates were clarified from sonicated nuclei and protein-DNA complexes were isolated with antibody.
ChIP was performed using HighCell ChIP kit following manufacturers protocol (Diagenode)
ChIP-Seq libraries were prepared using standard Illumina protocol. ChIP-enriched DNA samples (10-20 ng) were converted to blunt-ended fragments using a combination of 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 PE1.0/PE2.0 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 using the Bioanalyzer 2100 (Agilent) and each sample was sequenced in a single lane on the Illumina HiSeq 2000 (40 nucleotide read length).
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina Genome Analyzer II |
| |
|
| Description |
stable EED knockdown cells
|
| Data processing |
Illumina Casava1.7 software used for basecalling.
ChIP-seq reads were aligned to the hg19 genome assembly using bowtie with up to 3 mismatches
peaks were called using HPeak version 3 with the following setting: 25 bp window, 1e-3 significant level, using pair-end setting with dynamic fragment length
Genome_build: hg19
Supplementary_files_format_and_content: Tab-delimited text files were generated using HPeak ver. 3, which provided peak loci, height, width, summit location and log(p-value)
|
| |
|
| Submission date |
May 09, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Meng Zhao |
| E-mail(s) |
meng.zhao@emory.edu
|
| Phone |
404-712-8611
|
| Organization name |
Emory University
|
| Department |
BIOS
|
| Lab |
Steve Qin
|
| Street address |
1518 Clifton Rd, #369
|
| City |
Atlanta |
| State/province |
GA |
| ZIP/Postal code |
30322 |
| Country |
USA |
| |
|
| Platform ID |
GPL9115 |
| Series (1) |
| GSE42566 |
The Central Role of EED in the Orchestration of Polycomb Group Complexes |
|
| Relations |
| BioSample |
SAMN02141774 |
| SRA |
SRX276783 |
