Fig. 5

Additional validation of the detection of H3K9me3-H3K36me2/3 double modified chromatin. a Chromatin interacting domain precipitation (CIDOP) and ChIP quantified by real-time PCR (qPCR). Twelve regions (Additional file 1: Table S4) containing only H3K9me3, only H3K36me3 and both marks were selected. CIDOP was conducted with the D3PWWP-M8Chromo double domain and the two variants with one inactivated binding pocket D3PWWP-M8Chromo* (with inactivated H3K9me3 binding pocket) and D3PWWP*-M8Chromo (with inactivated H3K36me2/3 binding pocket). In addition, ChIP experiments with H3K9me3 and H3K36me3 antibodies were performed to confirm the presence or absence of both marks. Each experiment was conducted in two biological replicates. Error bars show the SEM. b Sequential CIDOP-ChIP experiments were conducted on H3K9me3 only, H3K36me3 only and double mark regions. First, H3K9me3-specific M8Chromo CIDOP was conducted followed by H3K36me3 ChIP. The pull-down was analyzed by semiquantitative PCR