The objective of this proposal is the development of methods and reagents for use in the study of deletion mutations at the molecular level. Such technology has not been developed for the analysis of deletion mutations which are induced by an important class of environmental mutagens/carcinogens and is necessary for the assessment of potential hazard to man from these compounds. the hypoxanthine guanine phosphoribosyltransferase (hprt) gene in human cells will be the molecular target. During Phase I, inverse polymerase chain reaction (PCR) technology will be applied to demonstrate its use in obtaining DNA sequence of breakpoint junctions and in developing unique probes (both PCR and hybridization) for analysis of deletion sites. In addition, PCR primer- pairs will be developed from yeast artificial chromosomes (YACs) which will allow assay of deletion size. These new reagents and methods will be used in Phase II studies for the molecular analysis of deletion mutations induced by chemical agents.
