Minimal extent of homology required for completion of meiotic recombination in Saccharomyces cerevisiae.

Abstract

The minimal length of contiguous homology required for successful completion of meiotic recombination was investigated by using heterologous insertions to delimit homologous segments of chromosome III in the yeast Saccharomyces cerevisiae. Constructs created in vitro by insertion of selectable markers into the LEU2 locus were transplaced into haploid strains, which were then mated to create diploids containing pairs of insertion heterologies at various distances. Analysis of the meiotic products from these diploids revealed a gradient in the frequency of both reciprocal and nonreciprocal recombination declining monotonically from the 5' end of LEU2. Both types of event were found to be restricted by the presence of the insertion heterologies. The spo13 single division meiosis was exploited to develop a plating assay in which LEU2 diploid spores containing reciprocally recombinant strands derived from events occurring completely within the interval flanked by the insertion heterologies were selected by random spore methods. Reciprocal recombination frequencies measured with this assay decreased linearly with extent, extrapolating to a minimal homology requirement of 150-250 nucleotides. When homology was most severely restricted, unexpected flanking marker configurations among reciprocal recombinants within LEU2 demonstrated the occurrence of complex recombination events. In addition to detecting reciprocal recombinants, the system is capable of measuring the probability that a non-reciprocal recombination event will have one end-point between the heterologous inserts and the other lying outside the interval. The minimal length of homology required for this aspect of recombination was found to be 25-60 nucleotides.