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Sample GSM1143123 Query DataSets for GSM1143123
Status Public on Feb 27, 2014
Title Med12_MCF7
Sample type SRA
 
Source name MCF-7_Med12_ChIP-seq
Organism Homo sapiens
Characteristics cell line: MCF-7
cell type: breast adenocarcinoma cells
chip antibody: Med12
chip antibody vendor: Bethyl
chip antibody cat. #: A300-774A
chip antibody lot #: A300-774A-1
Treatment protocol MCF7 cells were maintained in DMEM without phenol red and supplemented with 5% charcoal–dextran-treated FBS for 4 days prior to their use in estradiol (E2) treatment (10-7M, 30 min).
Growth protocol MCF7 cells were grown to 70% of confluence in DMEM with 10% FBS.
Extracted molecule genomic DNA
Extraction protocol For ChIP and FAIRE assays, DNA was extracted according to Svotelis et al. (Methods Mol Biol., 2009) and to Eeckhoute et al. (Genome Res., 2009) with few modifications. In brief, ~20-30.106 cells were cross-linked using 1.1% formaldehyde for 10 or 30 min at RT in 20ml of PBS 1X, reaction was quenched with 1ml of 2.5M Glycine. Cells were lysed with SDS lysis buffer (1% SDS, 10 mM EDTA, 50 mM Tris–HCI of pH 8.1) during 30 min at 4°C and sonicated to have DNA fragment size between 200 and 300 bp. For ChIP, lysate was incubated overnight at 4°C on a rotator with 5µg of antibody. 35 µl of protein-A magnetic beads (Invitrogen) were added during 2h before washing with TSE-150 (0.1% SDS, 1% Triton, 2 mM EDTA, 20 mM Tris–HCl of pH 8.1, 150 mM NaCl), TSE-500 (0.1% SDS, 1% Triton, 2 mM EDTA, 20 mM Tris of pH 8.1, 500 mM NaCl), LiCl detergent (0.25 M LiCl, 1% NP40, 1% sodium deoxycholate, 1 mM EDTA, 10 mM Tris of pH 8.1) and three times with TE 1X. For FAIRE, soluble chromatin was subjected to three consecutive phenol-chloroform extractions. DNA was then reverse-crosslinked overnight at 65ºC and treated with RNAse A for 30 min at 37°C, followed by Proteinase K for 2 h at 37°C. After purification, DNA was quantified with Picogreen (Invitrogen) before library preparation.
Libraries were prepared according to Illumina's instructions. Briefly, 10 ng (ChIP-seq) or 50 ng (FAIRE-seq) of DNA were end-repaired using a combination of T4 DNA polymerase and T4 polynucleotide kinase (Enzymatics). DNA fractions of ~300 bp were selectively isolated with solid-phase reversible immobilization (SPRI) beads (Agencourt AMPure, Beckman Coulter). After adapter ligation, DNA was PCR amplified with Illumina primers, the quality and quantity of each DNA library was analyzed using Agilent Bioanalyser before sequencing on HiSeq (Illumina) following the manufacturer's protocols.
 
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model Illumina HiSeq 2000
 
Description Sample 2
Data processing Reads were aligned to the Human Reference Genome (assembly hg18) using BWA version 0.6.1 (Li and Durbin, 2010).
Only sequence reads that were uniquely mapped to the genome and with a MAQ mapping quality score >10 were used for the analysis.
Aligned tags were converted to WIG files by F-Seq (Boyle et al. 2008)
Genome_build: hg18
Supplementary_files_format_and_content: wig files containing aligned tags
 
Submission date May 16, 2013
Last update date May 15, 2019
Contact name Nicolas Gévry
E-mail(s) nicolas.gevry@usherbrooke.ca
Organization name Université de Sherbrooke
Street address 2500 boulevard de l'université
City Sherbrooke
State/province Québec
ZIP/Postal code J1K 2R1
Country Canada
 
Platform ID GPL11154
Series (2)
GSE47027 LRH-1 governs vital transcriptional programs in endocrine sensitive and resistant breast cancer cells: LRH-1 ChIP-seq
GSE54892 LRH-1 governs vital transcriptional programs in endocrine sensitive and resistant breast cancer cells
Relations
BioSample SAMN02146463
SRA SRX278610

Supplementary file Size Download File type/resource
GSM1143123_Med12_MCF7.wig.gz 206.3 Mb (ftp)(http) WIG
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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