We propose using new molecular techniques to survey the spectrum of mutagenic events occurring at tk-l in mouse L5178Y cells. We will compare this spectrum with the range of abnormalities occurring in human cancer, to validate the mouse lymphoma assy (MLA) as an accurate reflection of events of human significance. Specifically, a large collection of mutants will be screened for tk-l allele loss by Southern blotting. Small lesions will be analyzed by manual and/or automated sequencing, following prescreening by single strand conformational polymorphism (SSCP) analysis to localize mutations and streamline the sequencing effort. The feasibility of SSCP analysis for screening mutants was demonstrated in Phase I. Phase II will provide full-scale use of this technique to streamline mutation analysis. We also propose developing a mouse chromosome Il painting probe, using a unique technology developed in our laboratory, preparative in situ hybridization. Such a probe will allow simplified analyses of chromosomal events in tk mutants. Primers and protocols employed in SSCP and sequence analysis may be made commercially available as kits or services. Painting probes will have utility in analysis of tk mutants, and in chromosome identification, and eventually form the basis for a new genetic toxicology assay for clastogens.

Project Start
1993-06-30
Project End
1995-06-29
Budget Start
1993-06-30
Budget End
1993-11-30
Support Year
Fiscal Year
1993
Total Cost
Indirect Cost
Name
Applied Genetics Laboratories, Inc.
Department
Type
DUNS #
City
Melbourne
State
FL
Country
United States
Zip Code
32904