The detection of K-ras mutations in colorectal cancer using the amplification-refractory mutation system

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Abstract
A total of 301 colorectal carcinoma (CRC) archival samples were analysed using the amplification-refractory mutation system (ARMS). Each sample was examined to determine the mutation status of codons 12 and 13 of the K-ras oncogene. The results from direct DNA sequence analysis carried out on 30 of the samples differed from the ARMS result in almost 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 polymerase chain reaction (PCR) was used to generate an amplicon from K-ras exon 1 from 23 of the samples. The PCR amplicons were cloned and sequenced, and the DNA sequence analysis of the cloned material was in agreement with the ARMS results in all but one case. This case represented a tumour that exhibited a five-nucleotide reversed inversion. The cloned sequence data confirm the sensitivity and specificity of the individual ARMS reactions and that it is possible in certain cases to detect additional, more complex, sequence variations.