Polymerase chain reaction‐based comprehensive procedure for the analysis of the mutation spectrum at the hypoxanthine‐guanine phosphoribosyltransferase locus in Chinese hamster cells

We have established a comprehensive procedure based on the polymerase chain reaction (PCR) to analyze the molecular spectrum of mutations at the hypoxanthine‐guanine phosphoribosyltransferase (hprt) locus in Chinese hamster cells. The procedure includes direct sequencing of PCR‐amplified hprt cDNA for locating point mutations in the expressed coding sequences, multiplex PCR amplification of the hprt exons for screening large deletions, and direct sequencing of PCR‐amplified hprt exons and their flanking regions for detecting intronic mutations resulting in mRNA splicing errors. Using this procedure, we have identified different types of mutations among a representative collection of spontaneous and induced HPRT‐deficient Chinese hamster cell mutants. This procedure is simple, rapid, accurate, and practical for a comprehensive study of the mutation spectrum in a large number of HPRT‐deficient Chinese hamster cell mutants.