DENATURING GRADIENT GEL-ELECTROPHORESIS (DGGE) ASSAY FOR K-RAS AND N-RAS GENES - DETECTION OF K-RAS POINT MUTATIONS IN HUMAN LUNG-TUMOR DNA

Point mutations in the ras oncogenes are very common in lung cancers a s well as in many of the other solid tumours. To effectively examine t he occurrence of these mutations in a large number of tumour samples, we have applied denaturing gradient gel electrophoresis (DGGE) for the analysis of point mutations of the K-ras and N-ras genes, using GC-cl amped, PCR-amplified DNA fragments. Among the 68 tumour DNA samples, w e detected 14 mutations in the K-ras gene. This was 78% of the mutatio ns identified by oligonucleotide hybridization. Altogether, eight of t he nine different kinds of base substitutions found in the tumour samp les were detected by the DGGE assay, representing substitutions at cod ons 12, 13, and 61 of the K-ras gene. Six of the detected mutations we re guanine to thymine transversions at codon 12; this was the most com mon type of alteration. On the basis of our experience, the present no n-radioactive DGGE analysis seems to be readily applicable for detecti on of the mutations in the K-ras and N-ras genes. Types of ras gene mu tations frequent in adenocarcinomas of the lung are also discussed.