Sample information curated by ChIP-Atlas

Antigen

Antigen Class
Input control
Antigen
Input control

Cell type

Cell type Class
Pluripotent stem cell
Cell type
ES cells
NA
NA

Attributes by original data submitter

Sample

source_name
mouse embryonic stem cells
day
0
treatment
None
presumed cell type
Embryonic stem cell
strain
C57BL/6J
antibody
No antibody, DNA input control

Sequenced DNA Library

library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Cells were lysed in nuclei extraction buffer (50 mM HEPES KOH (pH 7.5), 140 mM NaCl, 1m M EDTA, 10% glycerol, 0.5% NP-40 alternative, 0.25% Triton X-100) supplemented with protease inhibitor cocktail (Roche) for 10 minutes at 4 °C. Cell pellets were resuspended in protein extraction buffer (10 mM Tris-HCl (pH 8.0), 200 mM NaCl, 1 mM EDTA and 0.5 mM EGTA) supplemented with protease inhibitor cocktail and incubated for 10 minutes at room temperature. For sonication, pellets were resuspended in sonication buffer (10 mM Tris-HCl (pH 8.0), 0.5 mM EGTA, 1% Triton X-100, protease inhibitor cocktail) and then fragmented with a Bioruptor (Diagenode) sonicator at 4°C for 30 cycles using high amplitude and 30 seconds ON and 30 seconds OFF cycles to produce size ranges between 200 and 500 base pairs. 2 mg of each antibody was prebound by incubating with Protein A+G Dynabeads (Invitrogen 100-07D) in blocking buffer (PBS supplemented with 0.5% TWEEN) for 6 hours at 4°C. Washed beads were added to the chromatin lysate and incubated overnight. Samples were washed twice with low salt washing buffer (20 mM Tris-HCl (pH 8.0), 150 mM NaCl, 2 mM EDTA, 0.1% SDS, 1% Triton X-100), twice with high salt washing buffer (20 mM Tris-HCl (pH 8.0), 500 mM NaCl, 2 mM EDTA, 0.1% SDS, 1% Triton X-100) twice with LiCl buffer (10 mM Tris-HCl, 250mM LiCl, 1mM EDTA, 1% NP-40, 1% Na-deoxycholate), twice with TE (10 mM Tris-HCl pH 8.0, 1mM EDTA), supplemented with 50 mM NaCl and eluted in elution buffer (50 mM Tris-HCl, 10 mM EDTA and 1% SDS). Eluates were incubated at 65°C for 20 minutes, followed by de-crosslinking at 65°C for 6-15 hours. Samples were diluted in TE buffer and then treated with RNaseA (Roche) for 60 minutes at 37°C, followed by incubation with proteinase K (Sigma) for 45 minutes at 56°C. DNA was purified with phenol:chloroform:isoamyl alcohol. The following antibodies were used for ChIP experiments: anti-NCoR (ABE251, Millipore), anti-SMRT (ab24551, Abcam), anti-H3K27ac (ab4729, Abcam). Standard Illumina protocols

Sequencing Platform

instrument_model
Illumina HiSeq 2000

mm10

Number of total reads
24187917
Reads aligned (%)
96.9
Duplicates removed (%)
14.0
Number of peaks
583 (qval < 1E-05)

mm9

Number of total reads
24187917
Reads aligned (%)
96.6
Duplicates removed (%)
14.0
Number of peaks
642 (qval < 1E-05)

Base call quality data from DBCLS SRA