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| Status |
Public on Jun 12, 2018 |
| Title |
ERalpha s5 RAD1901 ChIPSeq |
| Sample type |
SRA |
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| Source name |
MCF7
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| Organism |
Homo sapiens |
| Characteristics |
cell line: MCF7
chip antibody: Estrogen Receptor alpha (Santa Cruz, sc-543, lot F1716)
cell type: breast cancer cell line
treatment: Rad1901
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| Treatment protocol |
Cells were treated with indicated compounds for 45 minutes before harvesting.
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| Growth protocol |
MCF cell were maintained in DMEM supplemented with 10% FBS, 4 mM L-glutamine and 1x non-essential amino acids. Cells were then washed 3 times in PBS and cultured for 72 hours in phenol red-free DMEM media supplemented with 10% charcoal-stripped serum prior to treatment.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Cells were formaldehyde fixed for 15 minutes (1% formaldehyde, 100 mM NaCl, 0.1 mM EDTA pH8.0, 5 mM Hepes pH 7.9). Fixation was stopped with 2.5M glycine and cells were washed 2X in PBS/0.5% Igepal CA-630. Cell pellets were snap frozen and chromatin was isolated by the addition of lysis buffer, followed by disruption with a Dounce homogenizer. Lysates were sonicated and the DNA sheared to an average length of 300-500 bp. Genomic DNA (Input) was prepared by treating aliquots of chromatin with RNase, proteinase K and heat for de-crosslinking, followed by ethanol precipitation.
Illumina sequencing libraries were prepared from the ChIP and Input DNAs by the standard consecutive enzymatic steps of end-polishing, dA-addition, and adaptor ligation. After a final PCR amplification step, the resulting DNA libraries were quantified and sequenced on Illumina’s NextSeq 500 (75 nt reads, single end).
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NextSeq 500 |
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| Data processing |
The 75-nt sequence reads are generated by Illumina sequencing (using NextSeq 500)
ChIP-seq reads were aligned to the hg19 genome assembly using BWA with default settings
Data were filtered using the following specifications: Only reads that pass Illumina’s purity filter, align with no more than 2 mismatches, and map uniquely to the genome are used in the subsequent analysis
peaks were called using macs2 version 2.1.1.20160309 with the following setting: Minimum FDR (q-value) cutoff -q (0.01), Effective genome size -g (hs), --bdg (True), --format (BAM), -c (pooled input is used as control)
treatment pileup bedgraph generated by macs2 is normalized to 10M totoal reads by dividing the number in "tags after filtering in treatment" field from the _peaks.xls file produced by macs2 times 10M
Genome_build: hg19
Supplementary_files_format_and_content: big wig files are converted from the normalized bedgraph file. The score respresents the normalized extended reads pileup signal
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| Submission date |
Jun 11, 2018 |
| Last update date |
Jun 12, 2018 |
| Contact name |
Zhenhua Wu |
| E-mail(s) |
zhenhuawu75@gmail.com
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| Phone |
6179592279
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| Organization name |
H3 Biomedicine
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| Street address |
300 Technology Square floor 5
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| City |
Cambridge |
| State/province |
MA |
| ZIP/Postal code |
02139 |
| Country |
USA |
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| Platform ID |
GPL18573 |
| Series (2) |
| GSE115607 |
ChIP-Seq analysis of estrogen deprived MCF7 cells treated with H3B-5942 and standards of care compounds |
| GSE115611 |
Discovery of Selective Estrogen Receptor Covalent Antagonists (SERCAs) for the treatment of ERa(WT) and ERa(MUT) breast cancer. |
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| Relations |
| BioSample |
SAMN09394290 |
| SRA |
SRX4193145 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM3184875_ERalpha_s5_RAD1901_summits.bed.gz |
214.6 Kb |
(ftp)(http) |
BED |
| GSM3184875_ERalpha_s5_RAD1901_treat_pileup_normalized10M.bw |
223.9 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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