Fig. 4
From: Alterations of p16-pRb Pathway and Chromosome Locus 9p21–22 in Sporadic Invasive Breast Carcinomas
Representative results of deletion mapping and microsatellite instability analysis with chromosome markers D9S162, D9S171, and IFNA and representative results of the PCR-based methylation assay. (A) M, marker (PUC19/Sau3 AI). Lanes 1–6, matched normal tumor samples (case 12) with polymorphism at the D9S162, D9S171, and IFNA loci and indication of loss of heterozygosity (LOH); (arrowhead), a1 and a2, alleles; Hd, heteroduplex lanes; LOH is shown with arrowhead). (B) Lanes 1 and 2, matched normal tumor samples (case 17) with no polymorphism at the D9S162 locus (Ho, homozygous), although the tumor sample showed MI, which is demonstrated as an aberrant band (α′); (arrow). Lanes 3 and 4, matched normal tumor samples with heterozygosity at D9S162 locus (case 33). α1 and α2, alleles; Hd, heteroduplex lanes. (C) De novo methylation of CDKN2/p16INK4a exon 1a in a ductal breast carcinoma (case 8). M, marker (PUC19/Sau3AI). Lanes 1 and 2, undigested PCR products from matched normal tumor sample. Lanes 3 and 4, digestion with methylation-sensitive enzyme HpaII revealed that CDKN2/p16INK4A in non-tumor tissue is unmethylated as no amplification by PCR is seen, whereas in the tumor, CDKN2/p16INK4A is methylated as digestion does not affect amplification. Lanes 5 and 6, digestion with non-methylation-sensitive enzyme MspI affects amplification in normal and tumor tissue.