Methylation analysis of PTTG1 expression. A) Western blotting analysis of each cell line cultured in the presence or absence of 5-Aza. B) MSP analysis of tumor cell lines. Converted DNA is subjected to three different PCR reactions. For each primer set, two primer pairs are used to determine if the PTTG1 locus is methylated (M) or unmethylated (U). Bisulfite conversion control is performed by amplifying p16-Ink locus. C) MSP analysis of tumor samples with low (TS1-3) or high PTTG1 expression levels (TS4-6).