HOT-SPOTS MUTATION ANALYSIS OF P53 GENE IN GASTROINTESTINAL CANCERS BY AMPLIFICATION OF NATURALLY-OCCURRING AND ARTIFICIALLY CREATED RESTRICTION SITES

We developed a rapid, simple method to detect ''hot spot'' point mutat ions of the p53 gene. A DNA fragment from cancer tissue of a surgical specimen was selectively amplified with specific oligonucleotide prime rs and then digested with restriction enzymes that recognized artifici al or naturally occurring restriction sites. To detect mutations in co dons 273 and 245, we created artificial Bst U1 and Bgl I sites by intr oducing a single nucleotide mismatch into the respective mutagenesis p rimers. We used the naturally occurring restriction sites of Msp I, Ha e III, and Hha I to detect mutations in codons 248, 249, and 175, resp ectively. In 74 cases of gastrointestinal cancer, 5 of 35 colorectal c ancers showed mutations; 1 of 15 cases of gastric cancers showed mutat ion; and 1 of 24 hepatocellular carcinomas showed mutation. This nonra dioactive method is an accurate and clinically useful way to detect ho t-spot point mutations of the p53 gene in gastrointestinal cancers.