|
| Status |
Public on Jan 23, 2015 |
| Title |
Spt16-10hr_ChIPSeq |
| Sample type |
SRA |
| |
|
| Source name |
embryonic stem cells
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6
cell type: embryonic stem cells
chip antibody: Spt16
|
| Treatment protocol |
Retinoic-acid differentiated tetON-Oct4 ESCs were treated with either PBS or doxycyline to induce expression of Oct4. These cells were fixed with formaldehyde, lysed using cell lysis buffer, sonicated to shear chromosomal DNA to 250-750 bp size, immunoprecipitated using antibodies against Oct4, Jmjd1c and Spt16.
|
| Growth protocol |
tet-ON Oct4 ESCs were maintained in ESC media containing 15% tetracycline-free FBS.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Lysates were clarified from sonicated nuclei and trancription factor-DNA complexes were isolated with antibody.
ChIP DNA libraries were prepared for sequencing using standard Illumina protocols
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina Genome Analyzer II |
| |
|
| Data processing |
Basecalls performed using CASAVA version 1.4
ChIP-seq reads were aligned to the NCBI Build 37 (UCSC mm9) of the mouse genome using Bowtie (Langmead et al., 2009). Duplicated reads were filtered (allowing no more than 2 reads at the same location).
Two programs were used to infer enriched binding regions (peaks): QuEST (Valouev et al., 2008) and MACS (Zhang et al., 2008). As negative control for defining background binding to the Rorγt analysis, we used an antibody for the isotype control immunoglobulin (IgG), and anti-RORγt antibody in a RORγt-deficient population. As a negative control for the Foxp3 analysis we used the Foxp3 antibody in EL4 cells (with no exogenous Rorc). As negative control for the EL4 peaks we used cells with no exogenous Rorc. The output of the two programs is a list of enriched binding regions (peaks) together with an estimate of statistical significance. For QuEST we retain all the peaks with p-value<10-8; for MACS we used a p-value cutoff of 10-8 or estimated FDR below 5%. We then applied a post-processing step, consolidating all the peaks that for a given algorithm (QuEST or MACS) was detected (up to a distance of 50bp) with all of the controls. We report this unified set of peaks. Associated p-values are minimum over QuEST or MACS.
Genome_build: mm9
Supplementary_files_format_and_content: TDF format; generated with igvtools count program
|
| |
|
| Submission date |
Jan 22, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Dean Tantin |
| E-mail(s) |
dean.tantin@path.utah.edu
|
| Phone |
(801) 587-3035
|
| Organization name |
University of Utah
|
| Department |
Department of Pathology
|
| Street address |
15 North Medical Drive East
|
| City |
Salt Lake City |
| State/province |
UT |
| ZIP/Postal code |
84112 |
| Country |
USA |
| |
|
| Platform ID |
GPL9250 |
| Series (1) |
| GSE65192 |
Pluripotency transcription factor Oct4 mediates stepwise nucleosome demethylation and depletion |
|
| Relations |
| BioSample |
SAMN03291007 |
| SRA |
SRX849881 |
