THE DETECTION OF K-RAS MUTATIONS IN COLORECTAL-CANCER USING THE AMPLIFICATION-REFRACTORY MUTATION SYSTEM

A total of 301 colorectal carcinoma (CRC) archival samples were analys ed using the amplification-refractory mutation system (ARMS). Each sam ple was examined to determine the mutation status of codons 12 and 13 of the K-ras oncogene. The results from direct DNA sequence analysis c arried out on 30 of the samples differed from the ARMS result in almos t 50% of the cases as a result of the relative excess of wild-type to mutated DNA sequences. To assess the validity of the ARMS data, the po lymerase chain reaction (PGR) was used to generate an amplicon from K- ras exon I from 23 of the samples. The PCR amplicons were cloned and s equenced, and the DNA sequence analysis of the cloned material was in agreement with the ARMS results in all but one case. This case represe nted a tumour that exhibited a five-nucleotide reversed inversion. The cloned sequence data confirm the sensitivity and specificity of the i ndividual ARMS reactions and that it is possible in certain cases to d etect additional, more complex, sequence variations.