Rapid diagnostic test for hereditary nonpolyposis colon cancer kindred using polymerase chain reaction

PURPOSE: We have identified a mutation in the hMLH1 gene from the proband of a hereditary nonpolyposis colorectal cancer kindred. We wished to develop a rapid test for this specific mutation to facilitate screening of other family members. METHOD: An allele-specific polymerase chain reaction strategy was used to detect a T insertion at the + 3 splice site post exon 9 in the hMLH1 gene. The test was evaluated on DNA in which the mutation status was known. RESULTS: A 130-base pair fragment was reliably amplified using the allele-specific polymerase chain reaction. The test is able to identify the mutant allele and to distinguish between normal, carriers (heterozygous), and tumor DNA samples. The mutant allele is not present in an unrelated hereditary nonpolyposis colorectal cancer cell line or in a sample of the normal population (n=49). CONCLUSIONS: This is a simple, rapid test that can determine carrier status in the members of a kindred at risk for this mutation. This mutation is unlikely to be a polymorphism. This test may now be evaluated in a clinical setting.