Sample information curated by ChIP-Atlas

Antigen

Antigen Class
TFs and others
Antigen
Spi1

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
Primary bone marrow-derived macrophages (BMDM)
strain
C57BL6/J
antibody
anti-PU.1 (SantaCruz, sc-352x)
treatment
no treatment

Sequenced DNA Library

library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Twenty million BMDMs per ChIP were crosslinked for 10 min at room temperature with 1% formaldehyde added in culture medium. Crosslink was stopped with ice-cold PBS containing 0.125M glycine for 5min. Nuclei were prepared by sequential incubation on ice for 5min in buffer A [10mM Tris-HCl (pH 8), 10mM EDTA, 0.25% Triton X-100], and for 30min in buffer B [10mM Tris-HCl (pH 8), 1mM EDTA, 200mM NaCl] (all buffers include proteases inhibitors). Nuclei were resuspended in sonication buffer [10mM Tris-HCl (pH 8), 1mM EDTA, 0.5% SDS, 0.5% Triton X-100, 0.05% NaDOC, 140mM NaCl] and sonicated with a Branson Digital Sonifier (Branson Ultrasonics) to an average size of 250bp. Sonicated chromatin was incubated overnight on a rotating platform at 4°C with a mixture of 20μl Protein A and 20μl Protein G Dynabeads (Invitrogen) pre-bound with 6μg of control IgG (sc-2028) or IRF8 (sc-6058), IRF1 (sc-640), PU.1 (sc-352) antibodies from Santa Cruz Biotechnologies or 3μg of H3 (ab1791), H3K27Ac (ab4729) from Abcam. Immune complexes were washed sequentially for 2min at room temperature with 1ml of the following buffers: Wash B [1% Triton X-100, 0.1% SDS, 150mM NaCl, 2mM EDTA, 20mM Tris-HCl pH 8]; Wash C [1% Triton X-100, 0.1% SDS, 500mM NaCl, 2mM EDTA, 20mM Tris-HCl pH 8), Wash D (1% NP-40, 250mM LiCl, 1mM EDTA, 10mM Tris-HCl pH 8]; and TEN buffer [50mM NaCl, 10mM Tris-HCl pH 8, 1mM EDTA]. After de-crosslinking by overnight incubation at 65°C, the DNA was purified with QIAquick PCR purification columns following manufacturer’s instructions (Qiagen). ChIP-seq libraries were prepared using Illumina TruSeq kit and sequenced on a HiSeq 2000 in paired-end 50bp configuration following manufacturer’s recommendations.

Sequencing Platform

instrument_model
Illumina HiSeq 2000

mm10

Number of total reads
72808103
Reads aligned (%)
98.5
Duplicates removed (%)
6.6
Number of peaks
115391 (qval < 1E-05)

mm9

Number of total reads
72808103
Reads aligned (%)
98.4
Duplicates removed (%)
7.0
Number of peaks
115355 (qval < 1E-05)

Base call quality data from DBCLS SRA