MiMB 262 - Genetic Recombination.pdf

METHODSMETHODS IN IN MOLECULAR BIOLOGYTMTM
Volume 262
ic
bination
ReviewsReviews andand ProtocolsProtocols
Edited by
Alan S. Waldman
Mitotic bination Rate Determination 3
1
Determination of Mitotic bination Rates
by Fluctuation Analysis in haromyces cerevisiae
Rachelle Miller Spell and Sue Jinks-Robertson
Summary
The study of bination in haromyces cerevisiae benefits from the availability of
assay systems that select for binants, allowing the study of spontaneous events that rep-
resent natural assaults on the genome. However, the rarity of such spontaneous bination
requires selection of events that occur over many generations in a cell culture, and the number
of binants increases exponentially following a bination event. To avoid inflation
of the average number of binants by jackpots arising from an event early in a culture, the
distribution of the number of binants in independent cultures (fluctuation analysis) must
be used to estimate the mean number of bination 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 bination 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 bination reporter system developed in
our lab to study the regulation of homeologous bination.
Key Words: fluctuation analysis, method of the median, confidence intervals, spontaneous
bination, mutation rate, inverted repeats, intron-based bination assay, homeologous
bination
1. Introduction
The study of DNA damage and subsequent repair by bination utilizes
systems that examine both spontaneous and induced damage. Although studies
of induced damage (by exogenous DNA-damaging agents or endogenous
expression of endonucleases) have the benefit of inflicting specific types of
damage, sometimes at known sites i