Sample information curated by ChIP-Atlas

Antigen

Antigen Class
TFs and others
Antigen
CTCF

Cell type

Cell type Class
Uterus
Cell type
SiHa
Primary Tissue
Cervix
Tissue Diagnosis
Carcinoma Squamous Cell

Attributes by original data submitter

Sample

source_name
SiHa
cell line
SiHa
chip antibody
CTCF
replicate
R3
sequencing type
Unpaired

Sequenced DNA Library

library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
CTCF ChIP-seq: We prepared 10 µl of both protein A and protein G beads through three washes of 5mg/mL Dulbecco's phosphate-buffered saline (dPBS) + bovine serum albumin (BSA). We added 10 µl of CTCF antibody (Cell Signalling Technology, Danvers, Massachusetts, Cat No. 2899, Lot 002) to the beads in 300 µl dPBS + BSA and left it to bind for > 6 hours of rotation at 4°C. After incubation, we washed the beads three more times with dPBS + BSA and resuspended in 100 µl of modified Radioimmunoprecipitation assay buffer (RIPA) (10 mmol/l Tris-HCl, pH 8.0; 1 mmol/l EDTA; 140 mmol/l NaCl; 1% Triton X-100; 0.1% SDS; 0.1% sodium deoxycholate) + protease inhibitor (PI). We collected 1 million cells by trypsinization and then fixed for 10 min at room temperature in 300 µl of dPBS + 1% formaldehyde. We added 15 µl of 2.5M glycine after fixing then washed the cells once in PBS + PI before resuspending them in 300 µl of modified RIPA + PI. We sonicated the samples for 32 cycles of 30 sec at full intensity using a Bioruptor Pico (Diagenode, Seraing, Belgium) and pelleted cell debris by spinning at 21130 xG for 15 min. We set aside 15 µl of the supernatant as an input control, and diluted the remaining supernatant with 1700 µl of modified RIPA + PI and 100 µl of washed beads. We incubated the samples at 4°C overnight with rotation. We washed the beads with the following cold buffers in order: modified RIPA, modified RIPA + 500 µmol l−1 NaCl, LiCl buffer (10 mmol l−1 Tris-HCl, pH 8.0; 1 mM EDTA; 250 mmol l−1 LiCl; 0.5% NP-40; 0.5% sodium deoxycholate), and finally twice with TE buffer (pH 8). We resuspended the samples and inputs in 100 µl of de-crosslinking buffer (1% SDS, 0.1000 mol l−1 NaHCO3) and incubated at 65°C for 6 h. We cleaned the samples and inputs using the Monarch PCR & DNA clean-up kit (New England BioLabs, Ipswich, Massachusetts), prepared libraries using the ThruPLEX DNA-seq Kit (Rubicon Genomics, Ann Arbor, Michigan), and size selected to 240 bp–360 bp using a PippinHT 2% Agarose Casette (Sage Science, Beverly, Massachusetts). We sequenced the samples to ∼25 million single-end 50 bp reads each at the Princess Margaret Genomics Core, Toronto, Ontario.

Sequencing Platform

instrument_model
Illumina HiSeq 2500

hg19

Number of total reads
25064551
Reads aligned (%)
80.6
Duplicates removed (%)
15.4
Number of peaks
21750 (qval < 1E-05)

hg38

Number of total reads
25064551
Reads aligned (%)
82.9
Duplicates removed (%)
13.7
Number of peaks
21903 (qval < 1E-05)

Base call quality data from DBCLS SRA