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Sample GSM4623478 Query DataSets for GSM4623478
Status Public on Feb 11, 2023
Title SMARCA4_786O_PBRM1KO_ChIPSeq_Rep1
Sample type SRA
 
Source name Cancer cell line
Organism Homo sapiens
Characteristics source: 786-O (ATCC CRL1932)
protein: SMARCA4
treatment: PBRM1_knockout
chip antibody: SMARCA4(AbCAM, ab110641, lot GR3208604-8)
Treatment protocol PBRM1 was knocked out using CRISPR-Cas9 mediated gene editing in 786-O, A498 and HK-2. Wildtype PBRM1 was cloned from normal kidney tissue. cDNA was either cloned into pbabe vector or pLenti-GFP.
Growth protocol Cells were grown in RPMI-1640 medium (Sigma) supplemented with 10% fetal bovine serum (Gibco). Cells were maintained in a 5% CO2-humidified incubator at 37°C.
Extracted molecule genomic DNA
Extraction protocol For each protein of interest, approximately 2x107 cells were cross-linked with 1% formaldehyde for 10 min at room temperature, and stopped by adding glycine to a final concentration of 0.2M. Chromatin was extracted and sonicated to ~500bp (Vibra cell, SONICS). The total volume of immunoprecipitation was 2 ml and the amount of antibody used was 20 µg. The input DNA was precleared with protein G Dynabeads (LifeTechnologies) for 2 hours at 4°C and then incubated with antibodies overnight at 4°C. Protein G beads were added the following day and mixture was nutated for 3 hours at 4°C. The beads were washed 6 times with wash buffer at room temperature.
At least 10 ng of the amplified DNA was used with NEBNext ChIP-Seq library prep reagent set (NEB). Each library was sequenced to an average depth of 30-50 million reads on HiSeq2500 or HiSeq4000.
 
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model Illumina HiSeq 2500
 
Data processing Sequencing reads were mapped against human genome reference hg19 using the Burrows-Wheeler Aligner (BWA) (version 0.6.2) 'mem' algorithm. Only reads with mapQ >10 and with duplicate removed by rmdup were used in the subsequent analysis
Significant peaks were called using MACS2 24 (q-value < 0.01).
Genome_build: hg19
Supplementary_files_format_and_content: ChIP-seq peaks are in bed format, including 5 columns: "chromosome" "start of region" "end of region" "peak name" "peak enrichment"
 
Submission date Jun 17, 2020
Last update date Feb 17, 2023
Contact name Xiaosai Yao
E-mail(s) xiaosai.yao@gmail.com, yao.xiaosai@gene.com
Phone +65 68088271
Organization name Genome Institute of Singapore
Street address 60 Biopolis Street, Genome, #02-01
City Singapore
ZIP/Postal code 138672
Country Singapore
 
Platform ID GPL16791
Series (2)
GSE152681 Genome-wide chromatin profiles of PBRM1 [ChIP-Seq]
GSE152735 Genome-wide chromatin profiles of PBRM1
Relations
BioSample SAMN15298806
SRA SRX15419260

Supplementary file Size Download File type/resource
GSM4623478_RCC365_peaks.bed.gz 1.3 Mb (ftp)(http) BED
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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