Determination of Mitotic Recombination Rates by Fluctuation Analysis in Saccharomyces cerevisiae

The study of recombination in Saccharomyces cerevisiae benefits from the availability of assay systems that select for recombinants, allowing the study of spontaneous events that represent natural assaults on the genome. However, the rarity of such spontaneous recombination requires selection of events that occur over many generations in a cell culture, and the number of recombinants increases exponentially following a recombination event. To avoid inflation of the average number of recombinants by jackpots arising from an event early in a culture, the distribution of the number of recombinants in independent cultures (fluctuation analysis) must be used to estimate the mean number of recombination events. Here we describe two statistical analyses (method of the median and the method of p₀ ) to estimate the true mean of the number of events to be used to calculate the recombination rate. The use of confidence intervals to depict the error in such experiments is also discussed. The application of these methods is illustrated using the intron-based inverted repeat recombination reporter system developed in our lab to study the regulation of homeologous recombination.