|
| Status |
Public on Jan 01, 2016 |
| Title |
THP1_115_Clock |
| Sample type |
SRA |
| |
|
| Source name |
THP-1 cells
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: THP-1
cell type: MLL-AF9 leukemia
chip antibody: anti-KAT13D/Clock (Abcam, ab3517, lot GR130170-5, 1mg/ml)
|
| Treatment protocol |
none
|
| Growth protocol |
Cells were grown in RPMI with addition of 50uM 2-mercaptoethanol and 10% fetal bovine serum.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Chromatin immunoprecipitation (ChIP) was performed as previously described (Kowalczyk, 2012) with minor modifications. Specifically, cells were first fixed in 2mM EGS (Thermo Scientific) for 30 minutes or 1 hour, followed by 1% of formaldehyde for 30 or 15 minutes at room temperature respectively. Chromatin was sonicated to a size less than 500bp and then incubated overnight with appropriate antibodies.
ChIP sequencing (ChIP-Seq) libraries were prepared through the process of DNA end-repair (Epicentre), A-base addition and adaptor ligation using indexed Illumina adaptors followed by enrichment PCR. All enzymatic steps are carried out using enzymes from New England Biolabs. Final libraries were pooled, size selected and sequenced on HiSeq2500 with 25 paired-end reads.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
Used in final analysis: no
ChIP conditions: 2mM EGS (Thermo Scientific) for 1 hour, followed by 1% of formaldehyde for 15 minutes at room temperature
|
| Data processing |
Aligned to genome; bowtie version 0.12.7 (bowtie -q -n 2 -e 70 --phred33-quals -X 1000 -p 4 -S -m 1[ …])
Merge SAMs from different runs; samtools version 1.2 (samtools merge […])
Sort BAMs by index; samtools version 1.2 (samtools sort […])
Remove PCR duplicates; picard ([…] REMOVE_DUPLICATES=true)
Call peaks using input as background; homer version 4.7.2 (findPeaks -style factor -fdr 0.01 -LP 0.01 -P 0.1 […])
Convert BAMs to BigWigs
Genome_build: hg19
Supplementary_files_format_and_content: *.bw: bigwig files containing genomic coverage; *.gff: GFF files containing peaks called for each factor
|
| |
|
| Submission date |
Jul 09, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Carl G de Boer |
| Organization name |
The Broad Institute
|
| Lab |
Aviv Regev
|
| Street address |
415 Main St
|
| City |
Cambridge |
| State/province |
MA |
| ZIP/Postal code |
02139 |
| Country |
USA |
| |
|
| Platform ID |
GPL16791 |
| Series (1) |
| GSE70686 |
ChIP-seq of circadian transcription factors in MLL-AF9 leukemia cell lines |
|
| Relations |
| BioSample |
SAMN03853103 |
| SRA |
SRX1091036 |
