Genome-wide Mapping of γ-H2A.X Marks

Chromatin immunoprecipitation using the anti-gamma H2A.X (Abcam, ab2893) was carried out according to a previously described protocol [43 (link)]. Libraries were constructed and bar-coded using TruSeq RS Cluster kit-HS (Illumina) and single-end sequencing (50 bp) was performed using an Illumina HiSeq-2500 sequencer at the Genomic Research Center of National Yang-Ming University (Taiwan) according to the manufacturer’s instruction. All reads were mapped to the human genome (hg38) using the bowtie2 alignment software [44 (link)]. The alignment results were used to call peaks by MACS [45 (link)]. The results of the peak signal were subject to the Integrative Genomics Viewer (IGV, Broad Institute) [46 (link)] for further comparison and analysis. We used UCSC hg38 and Rfam v11 to identify and annotate the ChIP-seq peak regions.

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Shih H.T., Chen W.Y., Wang H.Y., Chao T., Huang H.D., Chou C.H, & Chang Z.F. (2022). DNMT3b protects centromere integrity by restricting R-loop-mediated DNA damage. Cell Death & Disease, 13(6), 546.

Publication 2022

anti-gamma H2A.X HiSeq-2500 sequencer

Chip seq Chromatin immunoprecipitation Gamma Genomic H2a x Human Human genome

Corresponding Organization :

Other organizations : National Taiwan University, National Yang Ming Chiao Tung University, Chinese University of Hong Kong, Shenzhen

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Variable analysis

independent variables

Chromatin immunoprecipitation using the anti-gamma H2A.X (Abcam, ab2893)

dependent variables

ChIP-seq peak regions

control variables

Previously described protocol [43]
Library construction and bar-coding using TruSeq RS Cluster kit-HS (Illumina)
Single-end sequencing (50 bp) using an Illumina HiSeq-2500 sequencer
Alignment of reads to the human genome (hg38) using bowtie2
Peak calling using MACS
Visualization and analysis using Integrative Genomics Viewer (IGV)
Identification and annotation of ChIP-seq peak regions using UCSC hg38 and Rfam v11

positive controls

Not explicitly mentioned

negative controls

Not explicitly mentioned

Annotations

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