An integrated undergraduate laboratory exercise to demonstrate microbial evolution: petite mutants in Saccharomyces cerevisiae

Understanding that evolution progresses through generation of DNA variants followed by selection is a key learning outcome for biology students. We designed an integrated and innovative undergraduate laboratory exercise using Saccharomyces cerevisiae to demonstrate these principles. Students perform in vitro experimental evolution by repeatedly propagating large or small yeast colonies on a weekly basis. Small-colony variants known as petites arise by mutations that disrupt aerobic respiration. To demonstrate the effects of increased mutation rates, half of the selection lines are exposed to ultraviolet irradiation. To understand how the petite phenotype arises, polymerase chain reaction (PCR) is performed to examine mitochondrial DNA, while biochemical assays are used to assess the ability of petites to undergo aerobic respiration. This exercise demonstrates evolution by artificial selection over a suitably short timeframe and links the results to a critical biochemical process: the role of mitochondria in aerobic respiration and ATP production. By implementing these experiments, we successfully demonstrated that the frequencies of petite mutants in evolved populations varied according to the selection pressure we applied, and that petite mutants carried deletions in mitochondrial DNA as anticipated. Through an integrated learning context, this practical exercise promotes fundamental understanding of evolutionary processes and fosters critical thinking skills.