Figure 4

Bisulfite sequencing of cyclin D2 promoter region. The nucleotide sequence -1700 to -1250 bp from the transcription start site of the cyclin D2 gene promoter for the bisulfite sequencing analysis, and the relative regions of amplifications by MSP and bisulfite sequencing are shown. Genomic DNA from LnCap cells treated for 48 h was sodium bisulfite-treated, PCR amplified and subcloned. The sequencing results from 15-20 clones were used for analysis (upper figure, p values represent treatment values compared to vehicle), and representative 10 clones are shown in the dot plot. Each horizontal line represents the sequencing result of one subclone. Methylated CpG sites are shown as solid circles, whereas open circles indicate unmethylated CpG sites.