Sample information curated by ChIP-Atlas

Antigen

Antigen Class
TFs and others
Antigen
TP53

Cell type

Cell type Class
Digestive tract
Cell type
HCT 116
Primary Tissue
Colon
Tissue Diagnosis
Carcinoma

Attributes by original data submitter

Sample

source_name
HCT116
cell line
HCT116
chip antibody
P53 (FL393, sc-6243, Lot #B1709)
treatment
none

Sequenced DNA Library

library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
ChIP was performed using the ChIP-IT High Sensitivity (HS) Kit (Active Motif 53040) according to the manufacturer's recommended protocol with the following modifications as briefly described. Cells were cross-linked with 1% formaldehyde in DMEM culture medium for 10 min at room temperature, and quenched in 0.125 M glycine for 5 min. Cells were then washed twice with phosphate-buffered saline (PBS) followed by 3 times with lysis buffer (10 mM Tris-HCl pH 7.5, 10 mM NaCl, 3 mM MgCl2, 0.5% NP-40, 1 mM PMSF), and resuspended in ChIP Buffer (Active Motif 37516) supplemented with 1 mM PMSF and a proteinase inhibitor cocktail (Sigma-Aldrich P2714-1BTL). Chromatin was sheared by sonication (30 s alternated with 30-s cooling for 25 cycles) using a Bioruptor Pico ultrasonicator (Diagenode). Solubilised chromatin was used for immunoprecipitation by incubation overnight at 4 °C with 5 μg of the polyclonal antibody FL-393 against human p53 (Santa Cruz Biotechnology sc-6243). The immunoprecipitation reactions were transferred to Protein G Agarose Columns (Active Motif 53039) and incubated for 2 h at 4 °C. Then, each column was washed 4 times with 900 μl Wash Buffer AM1. The ChIPed samples were eluted using 100 μl Elution Buffer AM4 at 37 °C. Cross-linking was reversed by adding 5 μl Proteinase K (New England Biolabs P8107S) and incubated overnight at 65 °C. Input and ChIPed DNA were purified using the DNA purification columns. ChIP-seq libraries were prepared using MicroPlex Library Preparation Kit v2 (Diagenode C05010014) and the Agencourt AMPure XP system (Beckman Coulter A63880) was used to select 350–550 bp libraries. Sequencing was carried out on the Illumina MiSeq and HiSeq platforms.

Sequencing Platform

instrument_model
HiSeq X Ten

hg19

Number of total reads
39738987
Reads aligned (%)
60.8
Duplicates removed (%)
8.4
Number of peaks
454 (qval < 1E-05)

hg38

Number of total reads
39738987
Reads aligned (%)
62.0
Duplicates removed (%)
7.7
Number of peaks
814 (qval < 1E-05)

Base call quality data from DBCLS SRA