Sample information curated by ChIP-Atlas

Antigen

Antigen Class
TFs and others
Antigen
FOXA1

Cell type

Cell type Class
Breast
Cell type
MCF-7
Primary Tissue
Breast
Site of Extraction
Pleura
Tissue Diagnosis
Adenocarcinoma

Attributes by original data submitter

Sample

source_name
breast cancer MCF7 cells
tissue
breast cancer
cell line
MCF7L-P
genotype/variation
Parental
chip antibody
FOXA1 (abcam, ab23738, lot GR176970-2)

Sequenced DNA Library

library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Breast cancer cells were grown to 85% confluence in 15-cm plates and cross-linked with 1% formaldehyde (10 min) followed by quenching (125 mM glycine). Cells were washed in cold PBS and harvested by cell scraper in cold PBS with protease inhibitors (Roche). Cells were centrifuged at 3,220g for 5 min, snap-frozen, and stored at -80°C at 1×10^7 cells per pellet. Pellets were resuspended in cytosolic and then nuclear lysis buffer, and chromatin was sheared at 4°C using a water-bath sonicator (Bioruptor, Diagenode) for 20 min at high output (30 s on, 30 s off). Sonicated lysates were cleared by centrifuging at 20,000g for 10 min and diluted 4 times before pre-incubation with protein-A/G beads (Santa Cruz) for 30 min at 4°C. ChIP was performed by overnight incubation at 4°C with antibody against human FOXA1 (Abcam, ab23738), H3K27ac (Active Motif, #39134), or H3K4me1 (Abcam, ab176877), or Drosophila melanogaster histone variant H2Av (Active Motif, # 61752), followed by an additional 1 h incubation with protein-A/G beads. Beads were washed consecutively with low and high salt wash buffer, once with LiCl wash buffer, and once with TE buffer. DNA was eluted in elution buffer (50 mM NaHCO3 and 1% SDS) and then supplemented with 300 mM NaCl. Cross-links were reversed overnight at 67°C. RNA was digested at 37°C with 0.1 mg/ml RNase A for 30 min. DNA was purified with a PCR purification kit (Qiagen). Indexed libraries were prepared from ChIP DNA using the KAPA Hyper Library Preparation Kit (Kapa Biosystems). Libraries were amplified by 12 cycles of PCR, and then assessed for size distribution using the 4200 TapeStation High Sensitivity D1000 ScreenTape (Agilent Technologies) and quantified using the Qubit dsDNA HS Assay Kit (ThermoFisher). The indexed libraries were multiplexed, 10 libraries per pool. The pool was quantified by qPCR using the KAPA Library Quantification Kit (KAPA Biosystems) and then sequenced on the Illumina HiSeq 2000.

Sequencing Platform

instrument_model
Illumina HiSeq 2000

hg19

Number of total reads
54444643
Reads aligned (%)
90.2
Duplicates removed (%)
5.1
Number of peaks
18868 (qval < 1E-05)

hg38

Number of total reads
54444643
Reads aligned (%)
91.7
Duplicates removed (%)
4.2
Number of peaks
18543 (qval < 1E-05)

Base call quality data from DBCLS SRA