title: Molecular analysis of mutations in the human HPRT gene.
abstract: The HPRT assay uses incorporation of toxic nucleotide analogues to select for cells lacking the purine scavenger enzyme hypoxanthine-guanine phosphoribosyl transferase. A major advantage of this assay is the ability to isolate mutant cells and determine the molecular basis for their functional deficiency. Many types of analyses have been performed at this locus: the current protocol involves generation of a cDNA and multiplex PCR of each exon, including the intron/exon junctions, followed by direct sequencing of the products. This analysis detects point mutations, small deletions and insertions within the gene, mutations affecting RNA splicing, and products of illegitimate V(D)J recombination within the gene. Establishment of and comparisons with mutational spectra hold the promise of identifying exposures to mutation-inducing genotoxicants from their distinctive pattern of gene-specific DNA damage at this easily analyzed reporter gene.
authors.affiliation: Department of Environmental and Occupational Health, Graduate School of Public Health, University of Pittsburgh, 100 Technology Drive, Bridgeside Point, Pittsburgh, PA, 15219, USA, pho1@pitt.edu.
authors.forename: Phouthone
authors.initials: P
authors.lastname: Keohavong
authors.forename: Liqiang
authors.initials: L
authors.lastname: Xi
authors.forename: Stephen G
authors.initials: SG
authors.lastname: Grant
mesh_terms: D001483:Base Sequence; D002478:Cells, Cultured; D004252:DNA Mutational Analysis; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D005091:Exons; D017930:Genes, Reporter; D006801:Humans; D007041:Hypoxanthine Phosphoribosyltransferase / Q000235:genetics*; D060885:Multiplex Polymerase Chain Reaction; D009154:Mutation; D060152:V(D)J Recombination
chemical_list: D017931:DNA Primers; D018076:DNA, Complementary; D007041:Hypoxanthine Phosphoribosyltransferase