a, Western blot analysis of non-methylated CGI and bulk chromatin released from purified nuclei using antibodies against selected histone modifications and CpG-binding proteins (normalized to histone H3 levels). An antibody against histone H3 served as a loading control. Modifications associated with transcriptional activity, including H3 acetylated on amino acids K9 and K14, H3K4me3 and H3K4me2 were enriched in CGI chromatin, whereas the elongation and silencing marks H3K36me3, H3K9me3, H3K27me3 and H4K20me3 were depleted. The CXXC domain proteins Cfp1 and Kdm2a also showed enrichment within the CGI fraction. b, Cfp1 ChIP assayed by qPCR across the X-linked mouse Xist locus. Vertical strokes beneath the plot represent CpGs within the locus and the black bar above demarcates the CGI. The open box below the CpG map shows the region amplified for bisulphite analysis. IP, immunoprecipitation. c, Bisulphite analysis of input chromatin (female brain) and chromatin immunoprecipitated with Cfp1 antibodies and control MeCP2 antibodies. Twelve representative clones are shown from the total number sequenced (number in brackets). Solid and open circles represent methylated and non-methylated CpGs, respectively. Uncharacterized CpGs are represented as gaps.