Molecular analysis of two mouse dilute locus deletion mutations: Spontaneous dilute lethal^20J and radiation-induced dilute prenatal lethal Aa2 alleles

The dilute (d) coat color locus of mouse chromosome 9 has been identified by more than 200 spontaneous and mutagen-induced recessive mutations. With the advent of molecular probes for this locus, the molecular lesion associated with different dilute alleles can be recognized and precisely defined. In this study, two dilute mutations, dilute-lethal^20J (d^120J) and dilute prenatal lethal Aa2, have been examined. Using a dilute locus genomic probe in Southern blot analysis, we detected unique restriction fragments in d^120J and Aa2 DNA. Subsequent analysis of these fragments showed that they represented deletion breakpoint fusion fragments. DNA sequence analysis of each mutation-associated deletion breakpoint fusion fragment suggests that both genomic deletions were generated by nonhomologous recombination events. The spontaneous ^d120J mutation is caused by an interstitial deletion that removes a single coding exon of the dilute gene. The correlation between this discrete deletion and the expression of all dilute-associated phenotypes in d^120J homozygotes defines the d^120J mutation as a functional null allele of the dilute gene. The radiation-induced Aa2 allele is a multilocus deletion that, by complementation analysis, affects both the dilute locus and the proximal prenatal lethal-3 (pl-3) functional unit. Molecular analysis of the Aa2 deletion breakpoint fusion fragment has provided access to a previously undefined gene proximal to d. Initial characterization of this new gene suggests that it may represent the genetically defined pl-3 functional unit.