Detection of Loss of Heterozygosity in Tk-Deficient Mutants from L5178Y Tk +/- -3.7.2C Mouse Lymphoma Cells.

The mouse lymphoma assay (MLA), a forward mutation assay using the Tk-3.7.2C clone of the L5178Y mouse lymphoma cell line and the Thymidine kinase (Tk) gene, has been widely used as an in vitro genetic toxicity assay for more than four decades. The MLA can evaluate the ability of mutagens to induce a wide range of genetic events including both gene mutations and chromosomal mutations and has been recommended as one component of several genotoxicity test batteries. Tk-deficient mutants often exhibit chromosomal abnormalities involving the distal end of chromosome 11 where the Tk gene is located, in mice, and the type of chromosome alteration can be analyzed using a loss of heterozygosity (LOH) approach. LOH has been considered an important event in human tumorigenesis and can result from any of the following several mechanisms: large deletions, mitotic recombination, and chromosome loss. In this chapter, the authors describe the procedures for the detection of LOH in the Tk mutants from the MLA, and apply LOH analysis for understanding the types of genetic damage that is induced by individual chemicals.