|
| Status |
Public on Oct 04, 2016 |
| Title |
UND_RNAPII_B |
| Sample type |
SRA |
| |
|
| Source name |
UND_RNAPII
|
| Organism |
Homo sapiens |
| Characteristics |
immortalization type: SV40 large T antigen immortalized
cell type: human Fetal Osteoblasts (hFOB)
protocol: undifferentiated
chip antibody: Santa Cruz, sc-899, N-20
|
| Treatment protocol |
For induction of osteoblastic differentiation, the confluent cells were treated with differentiation cocktail as described previously (Karpiuk et al., 2012) and shifted to 39⁰C incubator.The undifferentiated cells were incubated with DMSO for 24 hr and then harvested. For differentiated state, cells were preatreated with JQ1 (250nM) or equal volume of DMSO one hour before the induction of differentiation. The media was refreshed every 48 hr with subsequent addition of inhibitor. The cells were differentiated for 5 days.
|
| Growth protocol |
Human fetal osteoblasts (hFOB) were maintained as previously described (Harris et al., 1995). In short, cells were maintained at 34⁰C in 5% CO2-atmosphere incubator in phenol red-free high-glucose Dulbecco’s modified Eagle’s media (DMEM/F12, Invitrogen) supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin (Sigma-Aldrich).
L3.6 cells were maintained in Minimum Essential Media (MEM, Invitrogen) supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin (Sigma-Aldrich) as previously described (Herreros-Villanueva et al., 2013).
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
The ChIP was performed with 20 min crosslinking in 1 % formaldehyde in PBS. Following chromatin shearing , 1-2µg antibody was used for immunoprecipitation overnight. The immunocomplexes were incubated 2 additional hours with protein A sepharose beads and subjected to washes. After reversing the cross-linking, the DNA was isolated by standart phenol-chloroform-isoamyl extraction procedure.
ChIP-Seq libraries were prepared with MicroPlex Library Preparation Kit following the manufacturer's instructions.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
processed data file: Merged_UND_RNAPII.bigwig
|
| Data processing |
The FASTQ files were mapped to the human reference genome (assembly hg19) using BOWTIE2 with very-sensitive presets in end-to-end mode on Galaxy Platform (version 2.2.6). The resulting BAM files of ChIPs were deduplicated with RmDup Samtools (Galaxy version 2.0). The corresponding BAM files of the replicates were merged within condition. The merged BAM files were normalized to fragments (reads) per kilobase per million (RPKM) using the bamCoverage tool on Deeptools Galaxy (version 1.1.4) with minimum mapping quality (MAPQ) set to 10 and with removal of duplicates (--ignoreDuplicates).
Genome_build: hg19
Supplementary_files_format_and_content: bigwig represent the normalized RPKM values of ChIP-seq signal
|
| |
|
| Submission date |
Jun 06, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Steven A. Johnsen |
| Organization name |
Robert Bosch Center for Tumor Diseases
|
| Lab |
Molecular Cancer Epigenetics
|
| Street address |
Auerbachstraße 112
|
| City |
Stuttgart |
| State/province |
BW |
| ZIP/Postal code |
70376 |
| Country |
Germany |
| |
|
| Platform ID |
GPL11154 |
| Series (2) |
| GSE82295 |
BRD4 localization to lineage-specific enhancers is associated with a distinct transcription factor repertoire [ChIP-Seq] |
| GSE82297 |
BRD4 localization to lineage-specific enhancers is associated with a distinct transcription factor repertoire |
|
| Relations |
| BioSample |
SAMN05210117 |
| SRA |
SRX1823807 |
