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| Status |
Public on Aug 04, 2022 |
| Title |
ChIP-seq_PolIISer2P_shMed26_2_E2_MCF7 |
| Sample type |
SRA |
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| Source name |
Human breast cancers
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| Organism |
Homo sapiens |
| Characteristics |
cell line: MCF7
treatment: shMed26_2_E2
chip antibody: PolIISer2p Abcam ab5095(Lot#GR3225147-1)
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| Treatment protocol |
For hormone treatments, cells were incubated at 37°C and 5% CO2 for at least 3 days in phenol red-free DMEM (GIBCO/Invitrogen) supplemented with 5% charcoal dextran-stripped FBS (GIBCO/Invitrogen). 17-β-Estradiol (E2; Steraloids, Inc.) was added to a final concentration of 100 nM. TNFalpha (R&D systems) was added to a final concentration of 20 ng/ml. 5α-dihydrotestosterone (DHT, Sigma) was added to a final concentration of 100 nM. The ethanol (EtOH) vehicle control was 0.05% in all samples.
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| Growth protocol |
MCF7 cells were maintained at 37°C and 5% CO2 in Dulbecco's modified Eagle's medium (DMEM, GIBCO/Invitrogen), supplemented with 10% fetal bovine serum (FBS, GIBCO/Invitrogen). LNCAP cells were maintained at 37°C and 5% CO2 in Advanced RPMI 1640 medium (GIBCO/Invitrogen), supplemented with 10% fetal bovine serum (FBS, GIBCO/Invitrogen).
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Briefly, cells were cross-linked with 1% formaldehyde at room temperature for 10 min. And then the cross-linking was quenched with 0.125M glycine for 5 min. Chromatin was fragmented using a Bioruptor® Pico (Diagenode) for 10 min at high power, with an interval of 30 s between pulses to get around 200bp fragments and precleared using 20 μl Protein G Dynabeads (Life Technologies, Cat# 10009D). Subsequently, the soluble chromatin was incubated with 2-5 μg antibodies at 4°C overnight. Immunoprecipitated complexes were collected using 30 μl Protein G Dynabeads, which have been saturated with PBS/1% BSA over night at 4°C, per reaction. For all ChIPs, after de-crosslinking overnight at 65°C, final ChIP DNA was extracted and purified using QIAquick spin columns (QIAGEN).
The libraries were constructed following Illumina’s Chip-Seq Sample prep kit.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 4000 |
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| Description |
PolIISer2P_shMed26_2_E2
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| Data processing |
Basecalls performed using CASAVA version 1.4
Seqencing reads were aligned to the hg38 genome assembly using Bowtie version 2.0.1.
Genome binding peaks were identified using the 'findPeaks' command in HOMER with setting of '-style factor': 200 bp peaks with 3-fold enrichment and 0.01 FDR significance over local tags.
To generate histograms for the average distribution of tag densities, position-corrected, normalized tags in 100 bp windows were tabulated within the indicated distance from specific sites in the genome. Clustering plots for normalized tag densities at each genomic region were generated using HOMER and then clustered using Gene Cluster 3.0.
Genome_build: hg38
Supplementary_files_format_and_content: bigWig
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| Submission date |
Aug 14, 2019 |
| Last update date |
Aug 04, 2022 |
| Contact name |
Yuliang Tan |
| E-mail(s) |
yut020@ucsd.edu
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| Organization name |
University of California, San Diego
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| Department |
School of Medicine
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| Street address |
Room 345, CMM West, 9500 Gilman Drive, Mail Code 0648
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| City |
La Jolla |
| State/province |
California |
| ZIP/Postal code |
92037-0648 |
| Country |
USA |
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| Platform ID |
GPL20301 |
| Series (2) |
| GSE135807 |
ChIP-seq assay in human cancer cells |
| GSE135808 |
Topoisomerase 1 Nicking-dependent HP1gamma Engagement Mediates Acute Activation of Phase-separated Enhancers |
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| Relations |
| BioSample |
SAMN12571273 |
| SRA |
SRX6712602 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4030524_PolIISer2P_shMed26_2_E2.ucsc.bigWig |
175.0 Mb |
(ftp)(http) |
BIGWIG |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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