A method to detect and characterize point mutations in transcribed genes: amplification and overexpression of the mutant c-Ki-ras allele in human tumor cells.
A significant percentage of human tumors contain activated ras oncogenes that have acquired oncogenic potential as a result of somatic point mutations at codon 12 or 61 of the encoded ras gene product. We report here a method to detect and characterize mutations in ras genes that is based on the ability of pancreatic ribonuclease (RNase A; EC 3.1.27.5) to cleave RNA heteroduplexes containing single-base mismatches. Using this method, we show that certain human tumor cells contain mutant c-Ki-ras genes, and we define the nature and position of these mutations. At the same time, we describe the presence and estimate the expression of both normal and mutant c-Ki-ras alleles in the same tumor cells. This method should be useful for the diagnostic detection and characterization of single point mutations in expressed genes.
E
E
Winter
F
F
Yamamoto
C
C
Almoguera
M
M
Perucho
D000483:Alleles; D005784:Gene Amplification; D006801:Humans; D009154:Mutation; D009369:Neoplasms / Q000235:genetics*; D009857:Oncogenes; D012313:RNA / Q000378:metabolism; D012259:Ribonuclease, Pancreatic / Q000494:pharmacology; D014158:Transcription, Genetic
D012313:RNA; D012259:Ribonuclease, Pancreatic