TABLE 3

Examples of independent experiments

QuestionExperimental approach
Is my favorite ysm mutant defective in meiosis (i.e., sporulation)?A ysm homozygous diploid is constructed, and the sporulation frequency of this diploid is compared to that of a congenic wild-type diploid.
Is the genetic lesion causing the sectoring phenotype (i.e., YAC loss) located within the YAC or a yeast nuclear gene?YSM+/ysm− heterozygous diploids are constructed, sporulated, and tetrads are dissected. The red-sectoring phenotype is analyzed in YAC-containing spores. If a particular ysm mutation is cis (within the YAC), then 100% of the YAC-containing spores derived from that mutant are expected to display the mutant red-sectoring phenotype. In contrast, if the ysm mutation is trans (within a yeast nuclear gene), then the mutation is expected to segregate 2:2 in meiosis. As a result, only 50% of the YAC-containing spores from this particular mutant are expected to display the mutant red-sectoring phenotype.
Are microtubules defective in my favorite ysm mutant?A ysm mutant haploid and isogenic wild-type strain are compared for growth sensitivity to the microtubule-destabilizing drug benomyl.
Does my favorite ysm mutant display a defect in DNA replication?Growth is compared between a ysm mutant haploid and isogenic wild-type strain on agar plates containing the DNA replication inhibitor hydroxyurea.
Is my favorite ysm mutant able to repair damaged DNA?Cell viability is compared between a ysm mutant haploid and isogenic wild-type strain on agar plates upon exposure to mutagens (e.g., ultraviolet light, methyl methanesulfonate).
Does my favorite ysm mutant have an elevated mutation frequency?The frequency of forward mutation at the CAN1 gene (i.e., production of canavanine-resistant cells) is compared in ysm mutant haploid and isogenic wild-type haploid cells.
Is the cell cycle progression of my favorite ysm mutant similar to that of the isogenic wild-type strain?Wild-type and mutant ysm cells are synchronized with α-factor. α-factor is removed, and cell cycle progression [as assessed by yeast cell bud size (Alberts et al. 2001)] is examined by microscopic observation over time.
Does my favorite ysm mutant have an abnormal cell morphology?The cell morphology and size of the ysm mutant and isogenic wild-type cells are compared by microscopy.