Methods for determining spontaneous mutation rates

Abstract

Spontaneous mutations arise as a result of cellular processes that act upon or damage DNA. Accurate determination of spontaneous mutation rates can contribute to our understanding of these processes and the enzymatic pathways that deal with them. The methods that are used to calculate mutation rates are based on the model for the expansion of mutant clones originally described by Luria and Delbrück (1943) and extended by Lea and Coulson (1949). The accurate determination of mutation rates depends on understanding the strengths and limitations of these methods and how to optimize a fluctuation assay for a given method. This chapter describes the proper design of a fluctuation assay, several of the methods used to calculate mutation rates, and ways to evaluate the results statistically.

Fig. 1

An illustration of the constant increase in the mutant fraction after a population reaches a size sufficiently large so that the accumulation of mutants is simply a function of population size. Luria’s conventions are followed {Luria, 1951 632 /id}): k = the generation numbered backwards from 0; N = the number of cells present at each generation; N_t = the final number of cells in the population; μ = the mutation rate per cell (assuming a synchronous population). At each generation there are N_t/2^k individuals that produce μN_t/2^k new mutations, which will produce a total of μN_t mutant progeny by the last generation.

Fig. 2

Spreadsheet method to solve transcendental equations by iteration. The example is the Lea-Coulson median estimator. Method: insert the experimentally determined median in A1; try various values of m at A3 to get the value at A4 close to 0.