Figure 4

The 5,6-dichloro-1-beta-D-ribofuranosylbenzimidizole (DRB)- stimulated introduction of H3K9me2 and H3K9me3 is partially associated with minichromosomes containing RNAPII. Wild-type SV40 minichromosomes were isolated from cells at 48 hr post-infection with or without treatment with 5,6-dichloro-1-beta-D-ribofuranosylbenzimidizole (DRB) from 24 to 48 hr post-infection. Intact Minichromosomes were subjected to an ISFIP ChIP analysis in which minichromosomes were first immune selected with antibody to RNAPII. The minichromosomes bound by antibody to RNAPII were sonicated and the soluble chromatin fraction subjected to a second ChIP with antibody to either H3K9me2 or H3K9me3. The percentage of the treated and untreated chromatin containing H3K9me2 and H3K9me3 was determined by real-time PCR using primers that recognize the early region of the genome and the fold increase resulting from treatment calculated.