Mutational analysis is a powerful experimental method to probe gene function. Gene deletions and mutations conferring loss of function or conditional lethality indicate if a gene is essential or not under a variety of experimental conditions. Point mutations can reveal information about function that is not possible from studies of the wild-type gene in vivo or the purified gene product in vitro. Here, we describe three strategies to mutagenize targeted regions of the yeast genome and show, with examples, the use of different genetic selection and screening methods to identify mutants based on phenotype.