The procedures used to identify and isolate mutants, referred to as genetic screens, depend on whether the experimental organism is haploid or diploid and, if the latter, whether the mutation is recessive or dominant. Genes that encode proteins essential for life are among the most interesting and important ones to study. Since phenotypic expression of a mutation in an essential gene leads to death of the individual, clever genetic screens are needed to isolate and maintain organisms with a lethal mutation.

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In haploid yeast cells, essential genes can be studied through the use of conditional mutations. Among the most common conditional mutations are temperature-sensitive mutations, which are useful in organisms, such as bacteria and lower eukaryotes, that can grow at a range of temperatures. For instance, a single missense mutation may cause the resulting mutant protein to have reduced thermal stability, such that the protein is fully functional at one temperature (e.g., 23 °C) but begins to denature and is thus inactive at another temperature (e.g., 36 °C), whereas the normal protein would be fully stable and functional at both temperatures. A temperature at which the mutant phenotype is observed is called nonpermissive; a permissive temperature is one at which the mutant phenotype is not observed even though the mutant allele is present. Thus mutant strains can be maintained at a permissive temperature and then subcultured at a nonpermissive temperature for analysis of the mutant phenotype.

An example of a particularly important screen for temperature-sensitive mutants in the yeast S. cerevisiae comes from the studies of L. H. Hartwell and colleagues in the late 1960s and early 1970s. They set out to identify genes important in regulation of the cell cycle (during which a cell synthesizes proteins, replicates its DNA, and then undergoes mitotic cell division). Exponential growth of a single yeast cell for 20–30 cell divisions forms a visible yeast colony on solid agar medium. Because mutants with a complete block in the cell cycle would not be able to form colonies, conditional mutants were required to study mutations that affect this basic cellular process. To screen for such mutants, the researchers first exposed yeast cells to mutagens and then identified mutant yeast cells that could grow normally at 23 °C, but could not form a colony when placed at 36 °C (Figure 6-6a).

Once temperature-sensitive mutants were isolated, further analysis revealed that some indeed were defective in cell division. In S. cerevisiae, cell division occurs through a budding process, and the size of the bud, which is easily visualized by light microscopy, indicates a cell’s position in the cell cycle. Each of the mutants that could not grow at 36 °C was examined by microscopy after several hours at the nonpermissive temperature. Examination of many different temperature-sensitive mutants revealed a set that exhibited a distinct block in the cell cycle. These mutants were therefore designated cdc (cell division cycle) mutants. Importantly, these yeast mutants did not simply fail to grow, as they might have if they carried a mutation affecting general cellular metabolism. Rather, at the nonpermissive temperature, the mutants grew normally for part of the cell cycle, but then arrested at a particular stage of the cell cycle, so that many cells at that stage were seen (Figure 6-6b). Most cdc mutations in yeast are recessive; that is, when haploid cdc strains are mated to wild-type haploids, the resulting heterozygous diploids are neither temperature sensitive nor defective in cell division.