Sample information curated by ChIP-Atlas

Antigen

Antigen Class
TFs and others
Antigen
ZNF366

Cell type

Cell type Class
Blood
Cell type
Dendritic Cells
MeSH Description
ANTIGEN-PRESENTING CELLS of dendritic cell morphology found in the LYMPH NODES and other lymphoid tissues.

Attributes by original data submitter

Sample

source_name
monocyte-derived dendritic cells
cell type
monocyte-derived dendritic cells
treatment
R848
treatment time
24h
chip antibody
DC-SCRIPT/ZNF366 (cat# AF4707, lot# CALX011009A, R&D Systems)

Sequenced DNA Library

library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Five million human moDCs were stimulated with R848, followed by cross-linking with 1% formaldehyde for 15 min at room temperature. The reaction was quenched with 0.125M glycine and washed at 4˚C with three buffers: 1) PBS; 2) buffer consisting of 0.2% Triton X-100, 0.2% NP-40, 1 mM EDTA pH 8, 0.5 mM EGTA pH 8, 10 mM Tris-HCl pH 7.5, 10 mM NaCl; and 3) buffer consisting of 0.15 M NaCl, 1 mM EDTA, 0.5 mM EGTA, 50 mM HEPES pH 7.6. Cells were then resuspended in a ChIP-incubation buffer (0.15% SDS, 1% Triton X-100, 150 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 20 mM HEPES, 1X protease inhibitor mixture [Roche], 0.1% BSA) and sheared using a Bioruptor Next Generation sonicator (Diagenode). Sonicated chromatin was centrifuged for 5 min at 4˚C and supernatant was snap-frozen and kept until ChIP. 10 ng of input or ChIP-enriched DNA were end-paired using T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase, New England Biolabs), and T4 polynucleotide kinase (New England Biolabs), followed by purification using the QIAquick PCR purification kit (Qiagen). Subsequently, DNA was dA-tailed using the Klenow fragment (3’ to 5’ exo minus, New England Biolabs), followed by purification using the MinElute Reaction Cleanup kit (Qiagen) DNA was ligated to multiplex NEXTflex adapters (Bioo Scientific). IP and input DNA were purified by the MinElute reaction Cleanup kit, and amplified by PCR using the KAPA HiFi HotStart ReadyMix PCR kit (KAPA Biosystems) with the following program: 45 sec at 98°C for initial denaturation, 15 sec at 98C, 30 sec at 65C, 30 sec at 72°C for 4 cycles, followed by 1 minute at 72°C for final extension. Removal of excess adaptors and selection of 300bp bands was done using 2% E-Gel SizeSelect Agarose Gels (Invitrogen). Adapter-modified DNA fragments were enriched by PCR using the KAPA HiFi HotStart ReadyMix PCR kit for 8-10 cycles with the aforementioned program. To get rid of the 120bp adapterdimer, the PCR product was purified using Ampure beads (Beckman Coulter)

Sequencing Platform

instrument_model
Illumina HiSeq 2000

hg19

Number of total reads
33923102
Reads aligned (%)
37.7
Duplicates removed (%)
61.4
Number of peaks
12912 (qval < 1E-05)

hg38

Number of total reads
33923102
Reads aligned (%)
38.6
Duplicates removed (%)
60.6
Number of peaks
12909 (qval < 1E-05)

Base call quality data from DBCLS SRA