Methylation of hMLH1 promoter correlates with the gene silencing with a region-specific manner in colorectal cancer - PubMed (original) (raw)

Figure 2

Methylation of hMLH1 promoter determined by COBRA. (A) DNA of C1a, RKO, Lovo, VACO432, VACO481 and VACO457 (lanes 1–6, respectively) was treated with NaHSO3, amplified by PCR, digested with _Bst_UI, and separated on a 2% agarose gel. The ratio of the digested fragments (91 and 287 bp) to the total of the digested and undigested fragment (378 bp) represents the per cent of methylation at _Bst_UI site. The per cents of methylation in C1a, RKO, Lovo, VACO432, VACO481 and VACO457 are 95, 95, 0, 98, 0 and 0%, respectively. M, markers. (B) DNA from normal mucosa (lanes 1 and 3) and tumour (lanes 2 and 4) of two colorectal cancer patients C61 (lanes 1 and 2) and C64 (lanes 3 and 4) was used for methylation analysis in the proximal region. C61 tumour shows no methylation and C64 tumour shows complete methylation at _Bst_UI site in hMLH1 promoter. M, markers. (C) DNA from normal mucosa (lanes 1, 2, 5, 6) and tumours (lanes 3, 4, 7, 8) of two colorectal cancer patients C61 (lanes 1–4) and C64 (lanes 5–8) was analysed for methylation in the distal region. The undigested products and _Bst_UI digested products are shown in lanes 1, 3, 5, 7 and lanes 2, 4, 6, 8, respectively. M, markers.