- The sequences of six different primer pairs designed against RASSF1A, the length of the amplicon, and the number of C↓CGG and CG↓CG sites for each primer pair are shown in the upper panel
- The lower panel shows the location of C↓CGG (solid line) and CG↓CG (broken line) cleavage sites in RASSF1A. The primer pairs were designed to target these C↓CGG and CG↓CG sites. Methylation of the cytosine in the CpG dinucleotide motif within the C↓CGG and CG↓CG sites protects against enzymatic restriction cutting by HpaII and BstUI, respectively, and enables formation of amplicons. Unmethylation results in digestion of the target and no formation of amplicons. The location of the initiation site by the codon ATG is show by an arrow
