Figure 1.

Schematic representation of bi-directional allele-specific PCR. The wild-type primer set is shown in green and the mutant primer set in pink. Three possible fragments can be formed depending upon substrate DNA zygosity, as well as enzyme system and cycling parameters. The green amplicon defines the allele-specific product formed when only the wild-type primers are extended and the pink amplicon defines the allele-specific product formed when only the mutant primers are extended. A heterozygote is identified by the presence of both PCR products. The target region amplified by non-allele-specific outer primers is shown in pale grey. Our selected enzyme system was inefficient at amplifying longer sequences and in the presence of competition from a smaller fragment insufficient target region was amplified for visualisation on a stained gel.