 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Apr 01, 2014 |
| Title |
ChIPseq_PolII_iSLK219 |
| Sample type |
SRA |
| |
|
| Source name |
Kidney epithelium
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Kidney epithelial cells
infection: rKSHV.219
cell line: iSLK-219
chip antibody: Pol II (Santa Cruz Biotechnology, sc-899 X)
|
| Treatment protocol |
When indicated, 1 µM doxycyclin (dox) and 500 µM phosphonoformic acid (PFA) were added to the culture medium for 72 hrs
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
1 x 10^7 cells were fixed with 1% formaldehyde 5 minutes prior to harvest. Nuclei preparations were done using the truChIP High Cell Chromatin Shearing Kit with SDS from Covaris and chromatin was sheared with a Covaris E220 Ultrasonicator. The average size of chromatin fragments was ~200 bp. Antibodies for ChIP were coupled to Dynabeads (Invitrogen) and incubated with chromatin extracts for 4 hours at 4C in cold RIPA buffer (10 mM Tris pH 8.0, 1 mM EDTA, 150 mM NaCl, 5% glycerol, 0.1% Na deoxycholate, 0.1% SDS, 1% Triton X-100, protease inhibitors (Complete Mini, Roche)). Antibodies used for ChIP: rat α-LANA (Advanced Biotechnologies, 13-210-100), α-H3K4me3 (Abcam, ab8580), α-H3K27me3 (Millipore, 07-449), α-H3K36me3 (Abcam, ab9050), and α-Pol II (Santa Cruz Biotechnology, sc-899 X). The immunoprecipitated chromatin was sequentially washed in high salt RIPA buffer (500 mM NaCl) and LiCl wash buffer (50 mM Tris pH 8.0, 1 mM EDTA, 250 mM LiCl, 1% NP-40, 0.5% Na deoxycholate). Chromatin crosslinks were reversed by incubating samples overnight at 65°C in the presence of proteinase K (200 µg/ml, Invitrogen). DNA was recovered using AMPure XP beads (Beckman Coulter, Inc).
Libraries were prepared according to Illumina's instructions (TruSeq ChIP Sample Prep Kit, part # 15034288)
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina Genome Analyzer II |
| |
|
| Data processing |
Basecalls performed using CASAVA version 1.8
ChIPseq: reads were aligned to the hg19 genome assembly using Bowtie2 (v2.1.0, default parameters)
ChIPseq: peaks were called using MACS 1.4.2 using the default parameters except when otherwise specified (see Samples section)
RNAseq: reads were aligned to the hg19 genome assembly using TopHat2 (v2.0.8b, default parameters)
RNAseq: gene expression analysis was performed with the Cufflinks/Cuffdiff package v2.1.1 using default parameters.
Genome_build: hg19
Supplementary_files_format_and_content: ChIPseq: peak calling output files from the MACS program (tab-delimited text format). Contain peak genomic location, tag abundance and peak score (p-value, FDR)
Supplementary_files_format_and_content: RNAseq: output files from Cufflinks and Cuffdiff (tab-delimited format) that include FPKM values, gene expression ratios (LEC219 vs LEC) and statistical analysis (scores) for all hg19-annotated genes
|
| |
|
| Submission date |
Mar 28, 2014 |
| Last update date |
Apr 01, 2014 |
| Contact name |
ALEXANDRE MERCIER |
| Organization name |
NOVARTIS INSTITUTES FOR BIOMEDICAL RESEARCH
|
| Department |
INFECTIOUS DISEASES
|
| Lab |
DON GANEM
|
| Street address |
5300 CHIRON WAY
|
| City |
EMERYVILLE |
| State/province |
CA |
| ZIP/Postal code |
94608 |
| Country |
USA |
| |
|
| Platform ID |
GPL9115 |
| Series (1) |
| GSE56144 |
Site-Specific Association with Host and Viral Chromatin by KSHV LANA and its Reversal during Lytic Reactivation |
|
| Relations |
| BioSample |
SAMN02711816 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1359506_ChIPseq_PolII_iSLK219_hg19.bed.gz |
209.5 Mb |
(ftp)(http) |
BED |
| Processed data provided as supplementary file |
| Raw data not provided for this record |
|
|
|
|
 |
