|
| Status |
Public on Feb 22, 2018 |
| Title |
IP for BRD1 replicate 2 |
| Sample type |
SRA |
| |
|
| Source name |
hES HUES64
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: HUES64
chip antibody: anti-BRD1 (Abcam, ab71877)
|
| Growth protocol |
HUES64 cells were cultured at 37° C in mTeSR 1 media (STEMCELL Technologies) on plates coated with Matrigel (Corning). Media was replaced daily and cells were passaged with 0.05 M EDTA/PBS.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
ChIP was performed in HUES64 cells using the Myers lab ChIP-seq protocol (http://myers.hudsonalpha.org/documents/Myers%20Lab%20ChIP-seq%20Protocol%20v041610.pdf) with the following modifications. Nuclei were sonicated using a BioRuptor (Diagenode) for 20 cycles (30 sec on/30 sec off) on high power. After centrifugation at 4°C for 15 min at 20,000 rcf., 5% of total chromatin was set aside for input and the remainder was incubated with 10 ug of anti-BRD1 (Abcam, ab71877) or anti-RING1B (Abcam, ab3832) overnight at 4° C. Chromatin/antibody complexes were captured by incubation with Protein A Dynabeads (Invitrogen) for 2 hr at 4° with rotation. After cross-link reversal, DNA was purified by phenol-chloroform exctraction and ethanol precipitation. Illumina ChIP-seq libraries were prepared using the NEBNext ChIP-seq library preparation kit (New England Biolabs) and sequenced as described above.
ChIP
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Data processing |
Raw FASTQ files were quality filtered using the FASTX-toolkit ‘fastq_quality_filter’ tool (http://hannonlab.cshl.edu/fastx_toolkit/) so that only reads containing a Qual score of ≥ 20 at least 80% of base positions were kept.
Filtered reads were aligned to the hg19 human genome annotation using Bowtie2 with default parameters (Langmead et al. 2009). Reads with a MAPQ ≥ 20 were kept and extended to an estimated fragment length of 150 bp using deepTools (Ramirez et al. 2014).
Peaks were called for each IP replicate using HOMER ‘findPeaks’ tool with a matched input as control. Considering the broad nature of peaks upon visual inspection, we set peak parameters so that the minimum peak size was 1000 bp and the minimum allowable distance between peaks was 2500 bp.
Common peaks from two biological replicates were identified using DiffBind (R Bioconductor) for each IP and these common peaks sets were used for downstream analyses.
Genome_build: hg19
Supplementary_files_format_and_content: bigWig files showing RPM normalized tag density
|
| |
|
| Submission date |
Sep 29, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Mitzi Kuroda |
| E-mail(s) |
mkuroda@genetics.med.harvard.edu
|
| Organization name |
Brigham & Women's Hospital
|
| Lab |
NRB168
|
| Street address |
77 Ave. Louis Pasteur
|
| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02115 |
| Country |
USA |
| |
|
| Platform ID |
GPL16791 |
| Series (1) |
| GSE104059 |
Bivalent complexes of PRC1 with orthologs of BRD4 and MOZ/MORF target developmental genes in Drosophila |
|
| Relations |
| BioSample |
SAMN07718383 |
| SRA |
SRX3230266 |
| SRA |
SRX3230266 |
