Antigen

Antigen Class

TFs and others

Antigen

MBD2

Cell type

Cell type Class

Liver

Cell type

Liver

MeSH Description

A large lobed glandular organ in the abdomen of vertebrates that is responsible for detoxification, metabolism, synthesis and storage of various substances.

Sample

source_name

NorHep cells-MBD2

tissue

liver

cell line

NorHep

Sequenced DNA Library

library_strategy

ChIP-Seq

library_source

GENOMIC

library_selection

ChIP

library_construction_protocol

ChIP (Chromatin immunoprecipitation) was performed using ChIP-IT Express Kit (Active motif) according to the manufacturer protocol. Chromatin was sheared to an average size ranging from 200–500bp using Covaris (Covaris, Inc) sonicator. Cells (2million) were fixed using 37% formaldehyde (CalBiochem) at a final concentration of 1% for 10min at room temperature. Incubation of 1/7 volume of 1M glycine for 5min terminated the fixation process. After sonification the samples were centrifuged for 10min at 40C at maximal speed. The supernatant (chromatin) was then immunoprecipitated with the indicated antibodies. DNA and RNA were extracted using AllPrep DNA/RNA/miRNA Universal Kit (Qiagen) according to the manufacturer's protocol. CHIP bisulfite sequencing libraries from the different samples (control and SAM treatment cells) were prepared following the Illumina ChIP-Seq library protocol. Bisulfite conversion of the immunoprecipitated DNA was performed after addition of methylated adaptors and before amplification. EZ DNA Methylation-Gold Kit (Zymo) was used for bisulfite conversion according to the manufacturer's protocol. CHIP sequencing libraries were prepared following the Illumina ChIP-Seq library protocol without bisulfite conversion. RNA (4 µg) of control and Mbd2 knock down cells was prepared for sequencing using the TruSeq RNA Sample Prep Kit (Illumina, San Diego, USA) following the manufacturer's protocol and sequenced in duplicate using Illumina HiSEQ2K platform (Illumina, San Diego, USA)(50bp pair-end reads); Target DNA fragments were captured using capture array we designed covering promoters in human genome. Sequencing was performed on the Illumina HiSEQ2K platform using a standard 50 cycle paired-end read sequencing protocol according to the manufacturer's recommendations.

Sequencing Platform

instrument_model

Illumina Genome Analyzer IIx

hg38

Number of total reads

47484521

Reads aligned (%)

4.3

Duplicates removed (%)

12.1

Number of peaks

266 (qval < 1E-05)

hg19

Number of total reads

47484521

Reads aligned (%)

4.1

Duplicates removed (%)

11.5

Number of peaks

220 (qval < 1E-05)