Sample information curated by ChIP-Atlas

Antigen

Antigen Class
Histone
Antigen
H3K4me2

Cell type

Cell type Class
Unclassified
Cell type
Unclassified
NA
NA

Attributes by original data submitter

Sample

source_name
20130911_H3K4me2
cells pointed by barcodes
mESC: 1-1152
mescs strain
V6.5 strain
mefs source
NA
eml source
NA

Sequenced DNA Library

library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
The cells were trypsinized and washed prior to encapsulation To minimize the abundance of barcode adaptors concatemers we use PacI restriction enzyme (R0547L, NEB, USA) immediately after ChIP washing steps and while the chromatin is still bound to the ChIP beads. PacI digest in between bound concatemers and in the middle of each adapter to form 30bp DNA fragments that can be easily filter out using simple size selection. Next, we elute the chromatin by adding 2X elution buffer, digesting RNA contaminates with Rnase (11119915001, Roche Diagnostics, USA) and removing the nucleosomes with ProtenaseK (P8102S, NEB, USA). To purify the DNA we use AMPure XP beads (A63880, Beckman Coulter, USA) and follow with 14 rounds of Single-Cell-PCR to amplify the labeled DNA. To reduce unspecific Illumina adapter ligation we first dephosphorylate all 5’ ends with Antarctic Phosphatase (M0289L, NEB, USA) and then use BciVi enzyme (R0596L, NEB, USA) to specifically cleave the labeled DNA, leaving an A overhang at the 5’ end of all DNA fragments with single cell adapters. After ligating Illumina adapters, we digest concatemers again using PacI and perform 14 additional rounds of Illumina-PCR (PfuUltra II Hotstart PCR Master Mix, 600850, Agilent Technologies, USA) before sequencing the library of reads.  Size selected for 200bp-1000bp

Sequencing Platform

instrument_model
Illumina HiSeq 2500

mm10

Number of total reads
14115644
Reads aligned (%)
32.5
Duplicates removed (%)
1.1
Number of peaks
11 (qval < 1E-05)

mm9

Number of total reads
14115644
Reads aligned (%)
32.2
Duplicates removed (%)
1.1
Number of peaks
11 (qval < 1E-05)

Base call quality data from DBCLS SRA