SCREENING FOR TP53 MUTATIONS IN OSTEOSARCOMAS USING CONSTANT DENATURANT GEL-ELECTROPHORESIS (CDGE)

We have previously developed conditions to screen for TP53 point mutat ions inside the conserved domains II-V of the gene by using constant d enaturant gel electrophoresis (CDGE). The present study reports condit ions for screening more of the codons in the frequently mutated region exon 5-8 and for detecting mutations in sequences encoding functional domains in the N- and C-terminal part of the protein. The ability of the CDGE technique to detect mutations was studied using controls with known sequence deviations. The resolution power of the technique to s eparate different types of mutations was tested by using seven differe nt single base pair mutants all residing in a stretch of four base pai rs. All mutants were separated from the wild type. The established CDG E screening strategy was then used to look for mutations in DNA from 2 8 osteosarcomas. Six (21.5%) of the samples were shown to have a TP53 mutation, and the exact characterization was performed by direct seque ncing. All of these were within the frequently reported mutated region exon 5-8. (C) 1993 Wiley-Liss, Inc.