The_role_of_CpG_island_methylation_in_immune_cell_gene_regulation. The_role_of_CpG_island_methylation_in_immune_cell_gene_regulation

Tissue-specific methylation patterns suggest a role for CpG island methylation in differentiation and cell-type-specific gene regulation. We have profiled CpG island methylation in different cells of the immune cell lineage to investigate this role. MBD-affinity purification combined with next generation sequencing was used to analyse CpG island methylation in dendritic cells, B cells, Th1, Th2 and naïve T cells. ChIP-seq was carried out to determine RNA polymerase II binding sites in these cell types and this was compared to the methylation profiles obtained. Protocol:Genomic DNA was extracted, sonicated and isolated by CxxC affinity purification (CAP) using recombinant CXXC from mMBD1 as described in Illingworth et al., PLoS Biology, 2008, or with MBD affinity purification (MAP) using recombinant MBD from hMeCP2 as described in Skene et al., Moll Cell 2010.Cells were purified by FACS or magnetic separation, crosslinked using 1% formaldehyde and chromatin sheared using sonication. Protein-DNA complexes were immunoprecipitated with the appropriate antibody and DNA was subsequently extracted. Adaptors were ligated to purified DNA which was then amplified by PCR. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Genome Analyzer following the manufacturer's protocols Sequencing was performed at the Wellcome Trust Sanger Institute; Hinxton, Cambridge, UK This data is part of a pre-publication release. For information on the proper use of pre-publication data shared by the Wellcome Trust Sanger Institute (including details of any publication moratoria), please see http://www.sanger.ac.uk/datasharing/