CHARACTERIZATION OF RAD4 GENE REQUIRED FOR ULTRAVIOLET-INDUCED EXCISION REPAIR OF Saccharomyces cerevisiae PROPAGATED IN Escherichia coli WITHOUT INACTIVATION

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Abstract
The previously isolated RAD4 gene designated as pPC1 from the genomic library of Saccharomyces cerevisiae (Yoon et al., 1985, Korean J. Genetics 7, 97-104) appeared to propagate in Escherichia coli and yet retained its complementing activitiy to rad4 mutants without inactivation. The subcloned RAD4 gene was found to be localized within a 2.5 kb DNA fragment flanking BglII and BamHI sites in the insert DNA, and was shown to have the same restriction map as a yeast chromosomal DNA, as determined by Southern hybridization. Tetrad analysis and pulse-field chromosome mapping have revealed that the cloned RAD4 gene can be mapped and integrated into the yeast chromosome V, the actual size of this gene. DNA-tRNA hybridization has shown that the isolated RAD4 gene did not contain a suppressor tRNA gene. These results have indicated that the pPC1 is a functional RAD4 gene playing a unique role involved in the nucleotide excision repair of yeast without any genetic change during amplification in E. coli.