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| Status |
Public on Sep 16, 2022 |
| Title |
HCT116-NUP93-mAC,ChIP-Seq for BRD4,UT, biological replicate 1 |
| Sample type |
SRA |
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| Source name |
colorectal cancer cells
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| Organism |
Homo sapiens |
| Characteristics |
cell line: HCT116
antibody: BRD4(Cat#: A301-985A100, RRID:AB_2620184)
treatment: untreated
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
HCT116 cells were crosslinked in 2 mM discuccinimidyl glutarate (DSG, CovaChem, 13301-100) for 30 min first, then in 1% formaldehyde for 15 min at room temperature. Except for NUP93, all other ChIP-Seq experiments used cells that were crosslinked by 1% formaldehyde for 10 min at room temperature. After quenching with 0.125M glycine, we collected cell pellets, and extracted the nuclei by using buffer LB1 [50 mM HEPES-KOH (pH 7.5), 140 mM NaCl, 1 mM EDTA (pH 8.0), 10% (v/v) glycerol, 0.5% NP-40, 0.25% Triton X-100 and 1×cocktail protease inhibitor], and then LB2 [10 mM Tris-HCl (pH 8.0), 200 mM NaCl, 1 mM EDTA (pH 8.0), 0.5 mM EGTA (pH 8.0) and 1×cocktail protease inhibitor]. After centrifuge, cell nuclei were suspended in buffer LB3 [10 mM Tris-HCl (pH 8.0), 100 mM NaCl, 1 mM EDTA (pH 8.0), 0.5 mM EGTA (pH 8.0), 0.1% Na-deoxycholate, 0.5% N-lauroyl sarcosine and 1×cocktail protease inhibitor], and fragmented using Q800R3 sonicator (QSONICA).Sheared chromatins were collected by centrifugation, and were incubated with appropriate antibodies at 4°C overnight. The next morning, the antibody-protein-chromatin complex was retrieved by adding 40ul Protein G Dynabeads (Thermo Fisher Scientific, 10004D). Immunoprecipitated DNA was de-crosslinked by 65ºC heating overnight, treated by proteinase-K, and were harvested by phenol chloroform or by Qiagen Quick DNA extraction kit.
ChIP-Seq library construction using NEBNext® Ultra™ II DNA Library Prep Kit for Illumina (NEB, E7645L).
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
NextSeq 550 |
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| Data processing |
FASTQ data were trimmed by Cutadapt and Trimmomatic.
PRO-Seq Reads were aligned using STAR with the hg19 build of the human genome. Read counts for Refseq mRNAs were determined using HTSeq-count.
Peaks for ChIP-Seq and Cut&Run were called by using MACS2.
HiC-Pro was used to process Hi-C and HiChIP data. The HiC contact matrix normalized by ICE method was converted into mcool by cooler.
Identification of significant chromatin contacts(loops interaction) from HiChIP data by using FitHiChIP.
Assembly: hg19
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| Submission date |
Mar 20, 2022 |
| Last update date |
Sep 16, 2022 |
| Contact name |
chuangye Qi |
| E-mail(s) |
[email protected]
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| Organization name |
Baylor College of Medicine
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| Department |
Huffington Center
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| Lab |
Zheng
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| Street address |
6450 E Cullen St
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| City |
HOUSTON |
| State/province |
Texas |
| ZIP/Postal code |
77030 |
| Country |
USA |
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| Platform ID |
GPL21697 |
| Series (1) |
| GSE165463 |
Human Core Nucleoporin Selectively Regulates Gene Transcription but is Dispensable for 3D Genome Organization |
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| Relations |
| BioSample |
SAMN26813729 |
| SRA |
SRX14524663 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM5962442_HCT116-NUP93-mAC_UT_BRD4_ChIP-Seq_Rep1.bigwig |
744.9 Mb |
(ftp)(http) |
BIGWIG |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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