|
| Status |
Public on Jul 07, 2020 |
| Title |
Rbpj_shEts1_3-2 |
| Sample type |
SRA |
| |
|
| Source name |
THP-6 T-ALL cell line
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: THP-6 T-ALLs
transduction: Ets1-silencing shRNA
disease: T lymphoblastic leukemia/lymphoma
cell type: T lymphocyte
antibody: Rbpj (Cell Signaling Technology, 5313)
|
| Treatment protocol |
THP-6 T-ALL cells were lentivirally transduced with pLKO.1-type shRNA for 2 days and then selected with puromycin for 2 days prior to harvesting.
|
| Growth protocol |
T-ALL cell lines were grown in RPMI 1640 (Invitrogen) supplemented with 10% fetal bovine serum (Hyclone or Gibco), 2 mM L-glutamine, 2-mercaptoethanol (0.0005% (v/v), Sigma), penicillin and streptomycin.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Cells were crosslinked for most samples by addition of 1% formaldehyde to media at 37 C for 10 minutes, followed by glycine quenching and 2x washing in PBS. For Ets1, Cells were crosslinked with 2 mM DSG and 1% formaldehyde. For H3K27ac, cells were crosslinked with 1% formaldehyde at room temperature. Nuclei were isolated by hypotonic buffer lysis, and chromatin was fragmented with a QSonica Q800 bath sonicator in 0.3% SDS-containing lysis buffer. Samples were diluted 1:3.3 and chromatin complexes containing the target of interest were isolated by immunoprecipitation, followed by reverse crosslinking and DNA purification by SPRI. For input samples, 1:8diluted sonicated chromatin (whole cell extract; wce), was added directly to the reverse crosslinking step without immunoprecipitation.
Purified ChIP or input DNA was end-repaired and A-tailed. Illumina barcoded adaptors were ligated and PCR amplified. Final library was isolated by double SPRI purification (sequential reverse and forward SPRI) and paired-end sequenced (38 bp x 2) on illumina NextSeq.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
NextSeq 550 |
| |
|
| Data processing |
ChIP-Seq: Reads were aligned to the hg19 genome assembly using bwa-aln and bwa-sampe (bwa version 0.7.12)
Genome_build: hg19
Supplementary_files_format_and_content: ChIP-Seq: Data were filtered to remove PCR duplicates and reads mapping to >2 genomic sites. Then bigwig display files were generated with deepTools bamCoverage and UCSC wigToBigWig, and bed peak files were generated with Homer findPeaks (“style -factor” for transcription factors and “style -histone” for H3K27ac).
|
| |
|
| Submission date |
Oct 07, 2019 |
| Last update date |
Jul 07, 2020 |
| Contact name |
Mark Yat-fung Chiang |
| E-mail(s) |
[email protected]
|
| Organization name |
University of Michigan
|
| Department |
Internal Medicine/Heme-onc
|
| Lab |
Room 2838
|
| Street address |
109 Zina Pitcher Place
|
| City |
Ann Arbor |
| State/province |
MI |
| ZIP/Postal code |
48019-2200 |
| Country |
USA |
| |
|
| Platform ID |
GPL21697 |
| Series (2) |
| GSE138516 |
Ets1 confers context-dependent activation of Notch signaling in T-cell leukemia [ChIP-Seq] |
| GSE138660 |
Ets1 confers context-dependent activation of Notch signaling in T-cell leukemia |
|
| Relations |
| BioSample |
SAMN12984373 |
| SRA |
SRX6958366 |
