Sample information curated by ChIP-Atlas

Antigen

Antigen Class
TFs and others
Antigen
EGR1

Cell type

Cell type Class
Blood
Cell type
Monocytes
MeSH Description
Large, phagocytic mononuclear leukocytes produced in the vertebrate BONE MARROW and released into the BLOOD; contain a large, oval or somewhat indented nucleus surrounded by voluminous cytoplasm and numerous organelles.

Attributes by original data submitter

Sample

Alias
E-MTAB-6305:BC276 Mono EGR1
Broker name
ArrayExpress
Description
Protocols: Peripheral blood mononucleated cells (PBMC) were obtained from healthy donor buffy coats (Etablissement Français du sang, Rungis, France) or CMML whole blood, separated on Pancoll (Pan-Biotech, Dutscher, Brumath, France). Peripheral blood CD14+ monocytes were sorted with magnetic beads and the AutoMacs system (Miltenyi Biotech, Paris, France). After sorting, monocytes were cultured overnight at 1 million cells/mL in RPMI 1640 Glutamax medium (ThermoFisher Scientific) supplemented with 10% heat inactivated fetal bovine serum (FBS, Lonza, Amboise, France), 1% penicillin/streptomycin and 2mM L-Glutamine (ThermoFisher Scientific). Cells were cross-linked with addition of 1% formaldehyde directly to the culture medium for 10 min at RT with agitation. Fixation was stopped by addition of 125mM glycin during 5 min at RT with agitation and samples were then washed 2 times in ice-cold PBS before addition of SDS lysis buffer (Millipore, 10uL per 1.106 cells) supplemented with 1% protease inhibitor cocktail (Active Motif). Samples were vortexed and incubated 15 min on ice before 10 min sonication at 40W (Covaris S220, Woodingdean, UK). The chromatin immunoprecipitation was carried out using ChIP-it express kit according to manufacturer's instruction (Active Motif) with a polyclonal anti-EGR1 antibody (sc-110, santa cruz bioechnology). Enriched DNA from ChIP and Input DNA fragments were end-repaired, extended with an 'A' base on the 3′end, ligated with indexed paired-end adaptors (NEXTflex, Bioo Scientific, Proteigene, Saint Marcel, France) using the Bravo Platform (Agilent, Les Ulis, France), size-selected after 4 cycles of PCR with AMPure XP beads (Beckman Coulter, Villepinte, France) and amplified by PCR for 10 cycles more.
ENA checklist
ERC000011
INSDC center alias
Inserm U1170
INSDC center name
Inserm U1170
INSDC first public
2019-03-04T17:03:23Z
INSDC last update
2017-12-18T17:12:36Z
INSDC status
public
SRA accession
ERS2059569
Sample Name
ERS2059569
Title
BC276 Mono EGR1
age
not available
cell_type
monocyte
disease
normal
genotype
wild type
growth condition
overnight RPMI 10 percent FBS
organism
Homo sapiens
organism part
blood
phenotype
CD14 positive
sex
female

Sequenced DNA Library

library_name
BC276 Mono EGR1_s
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Peripheral blood mononucleated cells (PBMC) were obtained from healthy donor buffy coats (Etablissement Français du sang, Rungis, France) or CMML whole blood, separated on Pancoll (Pan-Biotech, Dutscher, Brumath, France). Peripheral blood CD14+ monocytes were sorted with magnetic beads and the AutoMacs system (Miltenyi Biotech, Paris, France). After sorting, monocytes were cultured overnight at 1 million cells/mL in RPMI 1640 Glutamax medium (ThermoFisher Scientific) supplemented with 10% heat inactivated fetal bovine serum (FBS, Lonza, Amboise, France), 1% penicillin/streptomycin and 2mM L-Glutamine (ThermoFisher Scientific). Cells were cross-linked with addition of 1% formaldehyde directly to the culture medium for 10 min at RT with agitation. Fixation was stopped by addition of 125mM glycin during 5 min at RT with agitation and samples were then washed 2 times in ice-cold PBS before addition of SDS lysis buffer (Millipore, 10uL per 1.106 cells) supplemented with 1% protease inhibitor cocktail (Active Motif). Samples were vortexed and incubated 15 min on ice before 10 min sonication at 40W (Covaris S220, Woodingdean, UK). The chromatin immunoprecipitation was carried out using ChIP-it express kit according to manufacturer's instruction (Active Motif) with a polyclonal anti-EGR1 antibody (sc-110, santa cruz bioechnology). Enriched DNA from ChIP and Input DNA fragments were end-repaired, extended with an 'A' base on the 3′end, ligated with indexed paired-end adaptors (NEXTflex, Bioo Scientific, Proteigene, Saint Marcel, France) using the Bravo Platform (Agilent, Les Ulis, France), size-selected after 4 cycles of PCR with AMPure XP beads (Beckman Coulter, Villepinte, France) and amplified by PCR for 10 cycles more.

Sequencing Platform

instrument_model
Illumina HiSeq 2000

hg19

Number of total reads
55297432
Reads aligned (%)
18.8
Duplicates removed (%)
67.1
Number of peaks
1997 (qval < 1E-05)

hg38

Number of total reads
55297432
Reads aligned (%)
23.2
Duplicates removed (%)
63.2
Number of peaks
3322 (qval < 1E-05)

Base call quality data from DBCLS SRA