Sample information curated by ChIP-Atlas

Antigen

Antigen Class
TFs and others
Antigen
BRD4

Cell type

Cell type Class
Pancreas
Cell type
PANC-1
Tissue
Pancreas/Duct
Disease
Epithelioid Carcinoma

Attributes by original data submitter

Sample

Alias
E-MTAB-7034:Panc_BRD4_rep_2
Broker name
ArrayExpress
Description
Protocols: Briefly, cells were crosslinked with 1% formaldehyde for 20 minutes and quenched by Glycine (125mM final concentration). Upon cell scraping, nuclear pellets were prepared in and washed with the nuclear preparation buffer (150mM NaCl, 20 mM EDTA, 50 mM Tris-HCl (pH 7.5), 0.5% v/v NP-40, 1% v/v Triton-X-100, 20 mM NaF). Then samples were sonicated in sonication buffer (150mM NaCl, 20 mM EDTA, 50 mM Tris-HCl (pH 8), 1% v/v NP-40, 0.5% v/v Sodium deoxycholate, 20 mM NaF, 0.1% SDS) for 30 cycles (in L3.6pl) and 25 cycles (in BxPC-3 and Panc-1) using a Bioruptor Pico (Diagenode) and a cycle setting of 30s on and 30s off. Consequently, samples were precleared by incubation with 50% slurry of Sepharose 4B (GE Healthcare) and incubated with antibody overnight. Antibodies included 1µg of p63 (4A4) (sc-8431, Santa-Cruz), 1µg of H3K27ac (196-050, Diagenode), 2µg of BRD4 (C15410337, Diagenode), and 1µg of Rabbit IgG (C15410206, Diagenode). Sepharose beads with Protein A (for rabbit antibodies) or Protein G (for mouse antibodies, 4A4 p63) were added to samples and incubated for 2 hours followed by washes, de-crosslinking, and DNA extraction. Extraction of DNA was performed using Roti(R)-phenol/chloroform/Isoamylalcohol (Roth) extraction followed by precipitation in 100% Ethanol and washes with 70% Ethanol. Libraries for DNA from ChIP were made using the Microplex Library Preparation kit v2 (Diagenode) according to the manufacturer’s instructions.
ENA checklist
ERC000011
INSDC center alias
Section of Tumor Epigenetics, Department of General, Visceral and Pediatric Surgery, University Medical Center Gottingen, 37075 Gottingen, Germany
INSDC center name
Section of Tumor Epigenetics, Department of General, Visceral and Pediatric Surgery, University Medical Center Gottingen, 37075 Gottingen, Germany
INSDC first public
2018-12-18T17:02:52Z
INSDC last update
2018-07-20T14:51:18Z
INSDC status
public
SRA accession
ERS2619739
Sample Name
ERS2619739
Title
Panc_BRD4_rep_2
cell_line
Panc1
cell_type
epithelial cell
disease
pancreatic ductal adenocarcinoma
organism
Homo sapiens
organism part
pancreas

Sequenced DNA Library

library_name
Panc_BRD4_rep_2_s
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Briefly, cells were crosslinked with 1% formaldehyde for 20 minutes and quenched by Glycine (125mM final concentration). Upon cell scraping, nuclear pellets were prepared in and washed with the nuclear preparation buffer (150mM NaCl, 20 mM EDTA, 50 mM Tris-HCl (pH 7.5), 0.5% v/v NP-40, 1% v/v Triton-X-100, 20 mM NaF). Then samples were sonicated in sonication buffer (150mM NaCl, 20 mM EDTA, 50 mM Tris-HCl (pH 8), 1% v/v NP-40, 0.5% v/v Sodium deoxycholate, 20 mM NaF, 0.1% SDS) for 30 cycles (in L3.6pl) and 25 cycles (in BxPC-3 and Panc-1) using a Bioruptor Pico (Diagenode) and a cycle setting of 30s on and 30s off. Consequently, samples were precleared by incubation with 50% slurry of Sepharose 4B (GE Healthcare) and incubated with antibody overnight. Antibodies included 1µg of p63 (4A4) (sc-8431, Santa-Cruz), 1µg of H3K27ac (196-050, Diagenode), 2µg of BRD4 (C15410337, Diagenode), and 1µg of Rabbit IgG (C15410206, Diagenode). Sepharose beads with Protein A (for rabbit antibodies) or Protein G (for mouse antibodies, 4A4 p63) were added to samples and incubated for 2 hours followed by washes, de-crosslinking, and DNA extraction. Extraction of DNA was performed using Roti(R)-phenol/chloroform/Isoamylalcohol (Roth) extraction followed by precipitation in 100% Ethanol and washes with 70% Ethanol. Libraries for DNA from ChIP were made using the Microplex Library Preparation kit v2 (Diagenode) according to the manufacturer's instructions.

Sequencing Platform

instrument_model
Illumina HiSeq 4000

hg19

Number of total reads
16535063
Reads aligned (%)
41.0
Duplicates removed (%)
57.5
Number of peaks
325 (qval < 1E-05)

hg38

Number of total reads
16535063
Reads aligned (%)
43.4
Duplicates removed (%)
55.4
Number of peaks
305 (qval < 1E-05)

Base call quality data from DBCLS SRA