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

Environ Mol Mutagen. 1992;19(4):267-73. doi: 10.1002/em.2850190402.

Abstract

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.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Base Sequence
  • CHO Cells
  • Cricetinae
  • DNA
  • Exons
  • Hypoxanthine Phosphoribosyltransferase / genetics*
  • Molecular Sequence Data
  • Mutation*
  • Polymerase Chain Reaction / methods*

Substances

  • DNA
  • Hypoxanthine Phosphoribosyltransferase