Detection of single-base mutations in a mixed population of cells: a comparison of SSCP and direct sequencing

Single-base-pair changes in several genes have been shown to be important in carcinogenesis. We have compared the sensitivity of two commonly used techniques for detection of single-base-pair changes. Defined populations of cells were prepared by mixing wild-type Chinese hamster ovary cells with cells carrying a known mutation in the CTP synthetase gene. RNA was extracted and analyzed for the mutation by single-strand conformational polymorphism and direct sequence analysis of polymerase chain reaction products. We found that both techniques were able to detect the mutation when it was present in 25% of the cells, but that direct sequencing was slightly more sensitive and able to detect the mutation when it was present in only 20% of the population.