The Epigenome of Barley: Development and Optimisation of a ChIP-Seq Protocol to Investigate Histone Modifications in Barley

Post-translational covalent modifications of histones contribute to the partitioning of a genome into euchromatin and heterochromatin and are important in development, gene regulation and genome stability. Chromatin immunoprecipitation followed by Next Generation Sequencing (ChIP-seq) is a method that can reveal DNA sequences associated with various histone modifications. Here, a ChIP-seq protocol specific to whole barley seedlings has been developed to survey the epigenome of barley. Antibodies raised against four different histone epitopes (H3K4me3, H3K9me1, H3K27me3 and H3K56ac) are being utilised in this project to capture DNA sequences associated with modified histones. DNA was extracted from immunoprecipitates of chromatin isolated from ten day old whole barley cv. Morex seedlings. qPCR was used to assay the level of enrichment of DNA after the immunoselective process. DNA samples which were immunoselected by anti-H3K9me1 and anti-H3K56ac were Illumina sequenced. The reads were mapped to the barley cv. Morex reference genome and peaks of enrichment were detected by peak calling programs. The sequencing results are being used to explore the function of these histone modifications by searching for patterns relating modifications to genomic features such as gene expression level, genetic recombination rate, gene evolutionary rate and transposable element abundance.