Sample information curated by ChIP-Atlas

Antigen

Antigen Class
TFs and others
Antigen
TP53

Cell type

Cell type Class
Bone
Cell type
U2OS
Tissue
bone
Lineage
mesoderm
Description
osteosarcoma from the tibia of a 15 year old, J. Ponten and E. Saksela derived this line (originally 2T) in 1964 from a moderately differentiated sarcoma, viruses were not detected during co-cultivation with WI-38 cells or in CF tests against SV40, RSV or adenoviruses, mycoplasma contamination was detected and eliminated in 1972, (PMID: 6081590)

Attributes by original data submitter

Sample

source_name
U2OS cells
cell line
U2OS cells
tumor
osteosarcoma
Sex
female
age
15 years
tissue/cells
bone
chip antibody
p53 polyclonal Ab (Diagenode, C15410083)
agent
Irradiated with 10Gy (1Gy/min)

Sequenced DNA Library

library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
2x10^6 cells were inoculated on 10 cm2 dish 2 days before experiments. A two-step cross-linking procedure was applied for an efficient fixation of NFκB dimers with DNA (Tian 2012). Cells were washed twice with PBS, a solution of DSG (disuccinimidyl glutarate) in 1x PBS was added to the final concentration of 2 mM, and then cells were incubated for 45 minutes at RT with rotation. Next, cells were washed twice with PBS and the second fixation was performed using 1% formaldehyde for 10 min. at RT. For analysis of p53 binding, cells were fixed only with formaldehyde (as described above). Formaldehyde fixation was quenched by glycine (125 mM final concentration) and then nuclei were isolated using buffers and protocol from iDeal ChIP-seq Kit for Transcription Factors (Diagenode). Chromatin of nuclei re-suspended in 200 µl was sheared using Bioruptor® PLUS combined with the Bioruptor® Water cooler & Single Cycle Valve (at HIGH power setting) with 3 rounds of 5 cycles 30sec.ON/30sec.OFF; chromatin fragments with approximate length 100-600 bp were obtained. Chromatin immunoprecipitation was carried out using the iDeal ChIP-seq Kit for Transcription Factors (Diagenode) with 3µl/sample of anti-RelA(p65) polyclonal antibody (C15310256, Diagenode) or anti-p53 polyclonal antibody (C15410083, Diagenode), or without antibody (mock-IP), according to the manufacturer protocol. Immunoprecipitated DNA fragments and input DNA were sequenced using the HiSeq 1500 system with TruSeq workflow (Illumina)

Sequencing Platform

instrument_model
Illumina HiSeq 1500

hg19

Number of total reads
49082226
Reads aligned (%)
97.3
Duplicates removed (%)
10.4
Number of peaks
2379 (qval < 1E-05)

hg38

Number of total reads
49082226
Reads aligned (%)
98.1
Duplicates removed (%)
8.7
Number of peaks
2588 (qval < 1E-05)

Base call quality data from DBCLS SRA