ANALYSIS OF SPONTANEOUS FRAMESHIFT MUTATIONS IN REV1 AND REV1-1 STRAINS OF SACCHAROMYCES-CEREVISIAE

Frameshift mutations occur by a number of mechanisms. To better unders tand the nature of these mechanisms, we determined the DNA sequence ch anges of 232 independent, spontaneous frameshift mutations in the HIS4 gene of REV1 and rev1-1 strains of Saccharomyces cerevisiae. All fram eshift mutants were selected based on their ability to revert the +1 f rameshift mutation his4-38. DNA sequence information was recovered usi ng two approaches - the double-strand gap repair of plasmid pMP4, and the polymerase chain reaction (PCR). Using these techniques, saturated mutation spectra for the spontaneous reversion of his4-38 were genera ted. The most frequently occurring mutational events in both strains w ere -1 frameshifts, but +2 frameshifts, larger deletions, larger inser tions and more complex mutations were also observed. Between the REV1 and rev1-1 strains, we noticed a significant difference in the distrib ution of -1 frameshift mutations. In addition, while for -1 frameshift events there was no significant difference between the reversion spec tra determined by double-strand gap repair or PCR, there was a surpris ingly significant difference between the types of frameshift mutations recovered by double-strand gap repair (only -1 frameshifts and one +2 frameshift), and those recovered using PCR (-1 frameshifts, +2 frames hifts, larger deletions and insertions, and more complex mutations). T his difference may reflect a selectional mechanism inherent in double- strand break repair that avoids chromosomal sequences which include co mplex alterations.