Sample information curated by ChIP-Atlas

Antigen

Antigen Class
Histone
Antigen
H3K4me3

Cell type

Cell type Class
Blood
Cell type
GM12878
Tissue
blood
Lineage
mesoderm
Description
B-lymphocyte, lymphoblastoid, International HapMap Project - CEPH/Utah - European Caucasion, Epstein-Barr Virus

Attributes by original data submitter

Sample

source_name
GM12878 cell line
biomaterial_provider
CC
cell line
GM12878
antibody
Anti-trimethyl-Histone H3 (Lys4), Millipore 07-473
cell number
1000

Sequenced DNA Library

library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Sample containing 10^6 cells was used as the starting material and centrifuged at 1,600 g for 5 min and washed twice with 1 ml cold PBS. Cells were crosslinked by adding 1 ml freshly made 1% formaldehyde and incubating at room temperature for 5 min on a shaker. Crosslinking was promptly terminated by adding 50 µl of 2.5 M glycine and shaking for 5 min. The pellet was resuspended in 130 μl Covaris buffer (10 mM Tris-HCl, pH 8.1, 1 mM EDTA, 0.1% SDS and 1× protease inhibitor cocktail) and sonicated using the following conditions. Using Covaris M 220 (Covaris), the sample was sheared at 4 oC with 75 W peak incident power, 5% duty factor, and 200 cycles per burst for 12 min. The sheared sample was centrifuged in a 4 oC centrifuge at 14,000g for 10 min, and the supernatant was transferred to a new tube. Sheared chromatin was diluted with IP buffer (20 mM Tris-HCl, pH 8.0, 140 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 0.1% (w/v) sodium deoxycholate, 0.1% SDS, 1% (v/v) Triton X-100, with 1% freshly added PMSF and PIC) and aliquoted into 25 μl samples containing 250, 500, 1500, and 5,000 cell samples and stored at -80 oC. Library preparation was performed using the Swift Bio S2 library preparation kit (Swift Biosciences) using 40 μl of purified DNA. 2.5 μl of the low EDTA TE buffer in the 50 μl amplification cycle reaction mix was replaced with 2.5 μl of 20x EvaGreen, and amplification was terminated after samples saw an >3000 RFU increase (BioRad CFX Connect). Amplified DNA was then eluted into 7 μl low EDTA TE buffer where 2 μl could be used for qPCR analysis for preliminary quality control analysis, Kappa DNA quantification, and Tapestation fragment size analysis and the other 5 μl for library pooling. Libraries were pooled at 10 nM for sequencing by Illumina HiSeq 4000 with single-end 50 nt read. LIFE-ChIP-seq (Low-Input Fluidized-bed Enabled Chromatin Immunoprecipitation combined with sequencing), procedures are detailed in our paper

Sequencing Platform

instrument_model
Illumina HiSeq 4000

hg38

Number of total reads
16674115
Reads aligned (%)
80.9
Duplicates removed (%)
47.7
Number of peaks
4900 (qval < 1E-05)

hg19

Number of total reads
16674115
Reads aligned (%)
80.3
Duplicates removed (%)
48.6
Number of peaks
4972 (qval < 1E-05)

Base call quality data from DBCLS SRA