Sample information curated by ChIP-Atlas

Antigen

Antigen Class
TFs and others
Antigen
Nelfe

Cell type

Cell type Class
Blood
Cell type
Macrophages
MeSH Description
The relatively long-lived phagocytic cell of mammalian tissues that are derived from blood MONOCYTES. Main types are PERITONEAL MACROPHAGES; ALVEOLAR MACROPHAGES; HISTIOCYTES; KUPFFER CELLS of the liver; and OSTEOCLASTS. They may further differentiate within chronic inflammatory lesions to EPITHELIOID CELLS or may fuse to form FOREIGN BODY GIANT CELLS or LANGHANS GIANT CELLS. (from The Dictionary of Cell Biology, Lackie and Dow, 3rd ed.)

Attributes by original data submitter

Sample

source_name
Nelf1_LPS1h_BMMF
strain
C57BL/6
age
8-12 weeks
Sex
male
cell type
bone-marrow derived macrophages (BMMF)
treated with
10 ng/ml LPS for 1h
chip antibody
NELF-E (Proteintech, 10705-1-AP)

Sequenced DNA Library

library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
For GR (Santa Cruz Biotech, sc-1004X), BRD4 (Abcam, ab84776) and p65 (Santa Cruz Biotech, sc-372X) ChIP-seq, nuclei were sonicated with Covaris S220 sonicator according to manufacturer's instructions (130 μl shearing buffer, 200 cycles/burst, 120 sec, DF 10). For Pol 2 (Santa Cruz Biotech, sc-9001X) and NELF-E (Proteintech, 10705-1-AP) ChIP-seq, cells were formaldehyde cross-linked and nuclei were sonicated as above to obtain fragments in 150-500 bp range. Input DNA was prepared from sonicated material saved prior to IP. Lysates were cleared by centrifugation at 14,000 rpm, 20 min, 4°C, and then incubated with respective antibodies using 40 μl of 50% protein A/G PLUS agarose beads (for GR, BRD4 and Pol 2) or 60 μl of Dynabeads (Invitrogen) (for p65) per reaction at 4°C overnight. GR, BRD4 and Pol 2 IP beads were washed 4x with RIPA buffer and once with TE buffer. p65 IPs were washed 6x with modified RIPA buffer containing 100 mM LiCl on a magnetic stand and once with TE buffer+50 mM NaCl. Each reaction was then incubated in TE+0.5% SDS+200 μg/ml proteinase K (Invitrogen, 25530049) for 2 h at 55°C, followed by 6 h at 65°C to reverse crosslinks. DNA was purified using phenol-chloroform extraction and ethanol precipitation or using Qiagen PCR purification kit. The efficiency of ChIP was assessed by qPCR. The integrity and quality of DNA was evaluated with Bionalyzer 2100 (Agilent Technologies) The integrity and quality of DNA was evaluated with Bionalyzer 2100 (Agilent Technologies) before using 10 ng of material to prepare Illumina-compatible sequencing libraries using Illumina Truseq ChIP sample prep kit. Library preparation and sequencing was performed by Weill Cornell Epigenomics Core. Libraries were sequenced by a HiSeq 2500 (50 bp, single-end).

Sequencing Platform

instrument_model
Illumina HiSeq 2500

mm10

Number of total reads
38740851
Reads aligned (%)
98.5
Duplicates removed (%)
11.3
Number of peaks
523 (qval < 1E-05)

mm9

Number of total reads
38740851
Reads aligned (%)
98.3
Duplicates removed (%)
11.3
Number of peaks
572 (qval < 1E-05)

Base call quality data from DBCLS SRA