A DNA repair gene required for the incision of damaged DNA is essential for viability in Saccharomyces cerevisiae

Abstract

A diploid strain (RAD3/RAD3) of Saccharomyces cerevisiae was transformed with an integration plasmid containing an internal fragment of the cloned yeast RAD3 gene. Integration by homologous recombination inactivated one of the diploid RAD3 genes, creating a recessive mutation. This mutation is inferred to be lethal in haploid cells since sporulation of diploid transformants segregated two viable and two inviable spores per tetrad, while integration of plasmids containing one or the other end of the RAD3 gene resulted in diploid transformants that segregated normally--i.e., four viable spores in each tetrad. Evidence that integration of the internal fragment occurred specifically at one of the RAD3 genes in the diploid is provided by DNA . DNA hybridizations. In addition, transformation of a diploid strain heterozygous for the RAD3 gene (RAD3/rad3-2) (carrying a rad3 mutation that does not affect the viability of haploid cells) results in the rad- phenotype in half of the transformants, indicating that the RAD3 gene was inactivated in these cells.