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.
Yu
Y J
YJ
Department of Preventive Medicine and Community Health, University of Texas Medical Branch, Galveston 77555-1010.
Xu
Z
Z
Gibbs
R A
RA
Hsie
A W
AW
D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D006224:Cricetinae; D004247:DNA; D005091:Exons; D007041:Hypoxanthine Phosphoribosyltransferase / Q000235:genetics*; D008969:Molecular Sequence Data; D009154:Mutation; D016133:Polymerase Chain Reaction / Q000379:methods*
D004247:DNA; D007041:Hypoxanthine Phosphoribosyltransferase