title: Analysis of in vivo mutation in the Hprt and Tk genes of mouse lymphocytes.
abstract: Assays measuring mutant frequencies in endogenous reporter genes are used for identifying potentially genotoxic environmental agents and discovering phenotypes prone to genomic instability and diseases, such as cancer. Here, we describe methods for identifying mouse spleen lymphocytes with mutations in the endogenous X-linked hypoxanthine guanine phosphoribosyl transferase (Hprt) gene and the endogenous autosomal thymidine kinase (Tk) gene. The selective clonal expansion of mutant lymphocytes is based upon the phenotypic properties of HPRT- and TK-deficient cells. The same procedure can be utilized for quantifying Hprt mutations in most strains of mice (and, with minor changes, in other mammalian species), while mutations in the Tk gene can be determined only in transgenic mice that are heterozygous for inactivation of this gene. Expanded mutant clones can be further analyzed to classify the types of mutations in the Tk gene (small intragenic mutations vs. large chromosomal mutations) and to determine the nature of intragenic mutation in both the Hprt and Tk genes.
authors.affiliation: Division of Genetic and Reproductive Toxicology, U.S. FDA/NCTR, 3900 NCTR Road, HFT-120, Jefferson, AR, 72079, USA, Vasily.Dobrovolsky@fda.hhs.gov.
authors.forename: Vasily N
authors.initials: VN
authors.lastname: Dobrovolsky
authors.forename: Joseph G
authors.initials: JG
authors.lastname: Shaddock
authors.forename: Robert H
authors.initials: RH
authors.lastname: Heflich
mesh_terms: D000818:Animals; D001483:Base Sequence; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D004252:DNA Mutational Analysis; D017931:DNA Primers / Q000235:genetics; D005787:Gene Frequency; D007041:Hypoxanthine Phosphoribosyltransferase / Q000235:genetics*; D051379:Mice; D009154:Mutation; D013601:T-Lymphocytes / Q000502:physiology*; D013937:Thymidine Kinase / Q000235:genetics*
chemical_list: D017931:DNA Primers; D007041:Hypoxanthine Phosphoribosyltransferase; D013937:Thymidine Kinase