Sample information curated by ChIP-Atlas

Antigen

Antigen Class
TFs and others
Antigen
MYC

Cell type

Cell type Class
Blood
Cell type
P493-6
NA
NA

Attributes by original data submitter

Sample

source_name
P493-6
treatment
1uM KI-MS2-008
timepoint
24 hour
antibody manufacturer
abcam
antibody catalog
ab56
epitope
MYC
background
P4936_WCE_MS2_24HR_TC1

Sequenced DNA Library

library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Chromatin immunoprecipitation sequencing. ChIP-seq for c-Myc, Max, and H3K27ac was performed as in (Lin et al., 2012) with minor changes. P493-6 cells were grown to a density of 1 × 10^6 cells / mL prior to the start of time course. At each time point, pellets of 50 million cells were isolated 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 1,650g for 5 min and flash frozen and stored at –80°C at 5 × 106 cells per pellet. Pellets were resuspended in cytosolic and then nuclear lysis buffer, and DNA was sheared at 4 °C using a water-bath sonicator (Bioruptor, Diagenode) for 25 min at high output (30 s on, 30 s off) in 1 ml of sonication buffer supplemented with 0.5% SDS. Sonicated lysates were cleared by centrifuging at 20,000g for 10 min and incubated overnight end over end at 4 °C with magnetic beads prebound with antibody. Beads were washed three times with sonication buffer, one time with sonication buffer with 500 mM NaCl added, once with LiCl wash buffer (20 mM Tris pH 8.0, 1 mM EDTA, 250 mM LiCl, 0.5% NP-40, 0.5% sodium deoxycholate), and once with TE. DNA was eluted in elution buffer (50 mM Tris-HCl pH 8, 10 mM EDTA, and 1% SDS). Cross-links were reversed overnight at 65 °C. RNA and protein were digested with 0.2 mg/ml RNase A for 2 h followed by 0.2 mg/ml proteinase K for 1 hour. DNA was purified with phenol chloroform extraction and ethanol precipitation. Libraries for sequencing were prepared using the Rubicon Thruplex DNA-seq/FD library preparation kit. An input of 50 ng of DNA or less was used, and following ligation, libraries were amplified per the manufacturer's instructions. Amplified libraries were then size selected using AMPure beads (Agencourt AMPure XP) according to the manufacturer's instructions. Further size selection was performed using a 2% gel cassette in the Pippin Prep (SAGE Sciences) set to capture fragments of 200–700 bp in size. Libraries were multiplexed at equimolar ratios and run on a NextSeq 500 instrument (75-bp, single-end reads).

Sequencing Platform

instrument_model
NextSeq 500

hg19

Number of total reads
16674608
Reads aligned (%)
8.1
Duplicates removed (%)
89.2
Number of peaks
178 (qval < 1E-05)

hg38

Number of total reads
16674608
Reads aligned (%)
9.0
Duplicates removed (%)
88.2
Number of peaks
149 (qval < 1E-05)

Base call quality data from DBCLS SRA