engleski

The influence of heterologous insertions on homologous recombination in Saccharomyces cerevisiae

Recombination between closely related but diverged (homeologous) sequences has been extensively studied due to the profound biological and practical significance of this process. Less is known about possible effect(s) of the presence of blocks of complete heterology in one of the recombining molecules. Therefore we developed the experimental system in which we can follow the influence of insertions on recombination between homologues. Heterologous inserts were placed by standard genetic manipulations either in the recipient (yeast genome) or in the donor (plasmid molecule) of genetic information. Transformants were selected by complementation of mutation in the ura3 gene and further analyzed by Southern blotting, polymerase chain reaction (peR) and in some cases by replica plating. In addition to the wild type yeast strain, we also transformed several DNA-repair mutants. Our results demonstrate strong effect of heterologous insertions on homologous recombination by decreasing the efficiency of integration and changing the spectrum of recombination events observed. We also show that the products of the genes PMS1, MSH2 and SRS2, but not MSH3, are involved in this process. We propose a model for recombination between homologues containing insertion and discuss the significance of our results for overall genome stability and evolution.

Saccharomyces cerevisiae; homologous recombination; DNA insertions; integrative plasmids

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