|
| Status |
Public on Jul 22, 2019 |
| Title |
BRD4_ChIPSeq_IBET151_5000nM_MV4;11_rep1 |
| Sample type |
SRA |
| |
|
| Source name |
AML cells
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Biphenotypic B Myelomonocytic Leukemia
chip antibody: BRD4 (Bethyl labs #A301-985A)
|
| Treatment protocol |
Cells were seeded at 5000 cells/well in 96well plates, 1ul of I-BET151 was added per well from 100x DMSO stocks (final DMSO concentration: 1%) and cells were incubated for 72hrs. Cell viability was measured using Cell TiterGlo assay kit (Promega) following manufacturer instructions
Cells were seeded at 5000 cells/well in 96well plates, 1ul of I-BET151 was added per well from 100x DMSO stocks (final DMSO concentration: 1%) and cells were incubated for 72hrs. Cell viability was measured using Cell TiterGlo assay kit (Promega) following manufacturer instructions
|
| Growth protocol |
MV4;11 and K562 cells were grown in RPMI-1640 supplemented with 10% FBS, in a humidified incubator (37 °C, 5% CO2).
MV4;11 and K562 cells were grown in RPMI-1640 supplemented with 10% FBS, in a humidified incubator (37 °C, 5% CO2).
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
K562 and MV4;11 cells were treated with either DMSO or increasing concentration of I-BET151 for 6 hours at 37C. After treatment, cells were cross-linked with 1% formaldehyde for 10 min at room temperature, and cross-linking was stopped by the addition of 0.125 M glycine. Cells were then lysed using the SimpleChIP® Enzymatic Chromatin IP Kit (CST #9004) and DNA was digested to mostly mononucleosomes as recommended by the manufacturer
Library preparation of immunoprecipitated DNA was done using the NEBNext® ChIPSeq Library Prep Master Mix Set for Illumina® (NEB) and samples were sequenced in a 50bp single end run on the Illumina HiSeq-2000
K562 and MV4;11 cells were treated with either DMSO or increasing concentration of I-BET151 for 6 hours at 37C. After treatment, cells were cross-linked with 1% formaldehyde for 10 min at room temperature, and cross-linking was stopped by the addition of 0.125 M glycine. Cells were then lysed using the SimpleChIP® Enzymatic Chromatin IP Kit (CST #9004) and DNA was digested to mostly mononucleosomes as recommended by the manufacturer
Library preparation of immunoprecipitated DNA was done using the NEBNext® ChIPSeq Library Prep Master Mix Set for Illumina® (NEB) and samples were sequenced in a 50bp single end run on the Illumina HiSeq-2000
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
BRD4_MV411_ChIP_BRD4_151_C_5000nM_RPGC_inputSub.bw
MV411_ChIP_BRD4_151_C_5000nM.bed
|
| Data processing |
Quality check for raw sequencing reads was performed using FastQC
ChIP-seq and Chem-seq reads were aligned to the hg19 genome assembly using bwa with default parameters
Data were filtered for duplicated reads using samtools
Peaks were called using a MACS2 derived pipeline on two replicates for each sample using a combination of lenient and strick p-value largely inspired by the IDR workflow.
Bigwig profiles were created using deeptools normalized using the RPGC method with input subtracted
Genome_build: hg19
Supplementary_files_format_and_content: processed data files include signal files (biwig) and peak files (bed)
|
| |
|
| Submission date |
Oct 01, 2018 |
| Last update date |
Jul 22, 2019 |
| Contact name |
Pierre Khoueiry |
| Organization name |
American University of Beirut
|
| Department |
Biochemistry and Molecular Genetics
|
| Street address |
Bliss street
|
| City |
Beirut |
| ZIP/Postal code |
11-0236 |
| Country |
Lebanon |
| |
|
| Platform ID |
GPL11154 |
| Series (1) |
| GSE120715 |
Quantitative analysis of bi-modal binding of BET proteins at promoters predicts I-BET sensitivity |
|
| Relations |
| BioSample |
SAMN10150462 |
| SRA |
SRX4781511 |
