Provide Sequence Analysis of TK and HPRT Gene Mutations

We propose using new molecular techniques to survey the spectrum of mutagenic events occurring at tL5178Y cells. We will compare this spectrum with the range of abnormalities occurring in human cancemouse lymphoma assy (MLA) as an accurate reflection of events of human significance. Specifically,mutants will be screened for tk-l allele loss by Southern blotting. Small lesions will be analyzed bsequencing, following prescreening by single strand conformational polymorphism (SSCP) analysis to land streamline the sequencing effort. The feasibility of SSCP analysis for screening mutants was dePhase II will provide full-scale use of this technique to streamline mutation analysis. We also propchromosome Il painting probe, using a unique technology developed in our laboratory, preparative inSuch a probe will allow simplified analyses of chromosomal events in tk mutants. Primers and protocand sequence analysis may be made commercially available as kits or services. Painting probes will hof tk mutants, and in chromosome identification, and eventually form the basis for a new genetic toxclastogens.National Eye Institute (NEI)