|
| Status |
Public on May 30, 2010 |
| Title |
Stat3fl/fl FoxP3-GFP, CD4-Cre<STAT3 deficient>, H3K4me3_ChIPSeq |
| Sample type |
SRA |
| |
|
| Source name |
in vitro polarized Th17 T cells
|
| Organism |
Mus musculus |
| Characteristics |
strain: Stat3fl/fl FoxP3-GFP, CD4-Cre<STAT3 deficient>
cell type: STAT3-deficient-Th17 polarized T cells from naïve
passages: mouse primary T cells cultured in vitro for 3 days and restimulated with IL6
chip antibody: H3K4me3
antibody manufacturer: AbCam
antibody catalog number: 8580
|
| Treatment protocol |
After 3 day culture in vitro, cells were restimulated for 1h with IL-6 (10 ng/ml) and then processed for ChIP-Seq.
|
| Growth protocol |
Control (STAT3fl/fl) and STAT3-/- (CD4 Cre; STAT3fl/fl) naïve CD4+CD44-CD62L+ T cells were isolated and sorted on the FACSAria flow cytometer. Cells were cultured for 72h with platebound anti-CD3, anti-CD28 (5 mg/ml each), IL-6 (10 ng/ml) and TGF-b (2.5ng/ml) with blocking antibodies to IL-2, IL-4 and IFN-g (all at 10 mg/ml). Cells were restimulated for 1h with IL-6 (10 ng/ml) and then processed for ChIP-Seq.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Lysates were clarified from sonicated nuclei and protein-DNA complexes were isolated with antibody. Libraries were prepared according to Illumina's instructions. Briefly, DNA was end-repaired and the blunt, phosphorylated ends were treated with Taq polymerase and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with Illumina primers for 18 cycles and library fragments of ~250 bp (insert plus adaptor and PCR primer sequences) were band isolated from an agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Genome Analyzer following the manufacturer's protocols.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina Genome Analyzer |
| |
|
| Description |
Chromatin IP against H3K4me3
|
| Data processing |
Alignment: Sequence reads were obtained and mapped to the mouse mm9 genomes using the Illumina Genome Analyzer Pipeline. All reads mapping with two or fewer mismatches were retained and read starts were summed in sliding windows of 200 bp to create summary windows.
Peaks: Peak detection was performed with SICER (Zang et al, 2009 Bioinformatics, 25:1952-1958)
|
| |
|
| Submission date |
May 05, 2010 |
| Last update date |
May 15, 2019 |
| Contact name |
Golnaz Vahedi |
| Organization name |
National Institutes of Health
|
| Department |
NIAMS
|
| Lab |
Lymphocyte Cell Biology Section
|
| Street address |
9000 Rockville Pike Bldg 10 Rm 13C101A
|
| City |
Bethesda |
| State/province |
MD |
| ZIP/Postal code |
20892 |
| Country |
USA |
| |
|
| Platform ID |
GPL9185 |
| Series (2) |
| GSE21669 |
Diverse Targets of the Transcription Factor STAT3 Contribute to T Cell Pathogenicity and Homeostasis [ChIP-seq] |
| GSE21671 |
Diverse Targets of the Transcription Factor STAT3 Contribute to T Cell Pathogenicity and Homeostasis |
|
| Relations |
| SRA |
SRX020336 |
| BioSample |
SAMN00013397 |
| Named Annotation |
GSM540721_H3K4me3Stat3KOTh17.bed.gz |
