Comprehensive TP53-denaturing gradient gel electrophoresis mutation detection assay also applicable to archival paraffin-embedded tissue

A comprehensive mutation detection assay is described for the entire coding region and all splice site junctions of TP53. The assay is based on denatu ring gradient gel electrophoresis, which follows either multiplex polymeras e chain reaction (PCR) applied to DNA extracted from fresh or frozen tissue samples or nested PCR applied to DNA extracted from paraffin-embedded tiss ue samples. in both instances, the analysis can be performed under a single set of conditions. when testing the assay on DNA from cultured lung cancer cell lines and from paraffin-embedded Dukes C colorectal carcinomas, signi ficant TP53 mutations were observed at high frequencies in 15 of 16 lung ca ncer cell lines (94%) and in 21 of 30 paraffin-embedded tissue samples of D ukes C colorectal carcinomas (70%). A substantial proportion of these signi ficant mutations occurred outside the evolutionary conserved region of TP53 in 4 of 16 lung cancer cell lines (25%) and in 11 of 30 paraffin-embedded colorectal carcinomas (37%). This underscores the importance of a comprehen sive TP53 mutation analysis in those instances that TP53 mutation is taken into account for diagnostic and prognostic purposes.