Project description:microarray experiment to test the gene expression in long term lines of mutator and non-mutator yeast. Here we use an experimental evolution approach to investigate the conditions required for evolution of a reduction in mutation rate and the mechanisms by which populations tolerate the accumulation of deleterious mutations. We find that after ~6700 generations four out of eight experimental mutator lines had evolved a decreased mutation rate.
Project description:microarray experiment to test the gene expression in long term lines of mutator and non-mutator yeast. Here we use an experimental evolution approach to investigate the conditions required for evolution of a reduction in mutation rate and the mechanisms by which populations tolerate the accumulation of deleterious mutations. We find that after ~6700 generations four out of eight experimental mutator lines had evolved a decreased mutation rate. 2 condition experiment, derived experimental evolution strains compared to their ancestor strain. We compared the expression profile of one of the mutator lines (m8) after 6700 generations with its mutator ancestor, and as a control, an evolved non mutator after 6700 generations was compared to to its non-mutator ancestor. In order to prepare cells for expression microarray, glass tubes containing 3 ml of YPD were inoculated from overnight cultures, and grown until the OD600 was approximately 0.3.
Project description:Industrial bioethanol production may involve a low pH environment,improving the tolerance of S. cerevisiae to a low pH environment caused by inorganic acids may be of industrial importance to control bacterial contamination, increase ethanol yield and reduce production cost. Through analysis the transcriptomic data of Saccharomyces cerevisiae with different ploidy under low pH stress, we hope to find the tolerance mechanism of Saccharomyces cerevisiae to low pH.
Project description:Population adaptation to strong selection can occur through the sequential or parallel accumulation of competing beneficial mutations. The dynamics, diversity and rate of fixation of beneficial mutations within and between populations are still poorly understood. To study how the mutational landscape varies across populations during adaptation, we performed experimental evolution on seven parallel populations of Saccharomyces cerevisiae continuously cultured in limiting sulfate medium. By combining qPCR, array CGH, restriction digestion and CHEF gels, and whole genome sequencing, we followed the trajectory of evolution to determine the identity and fate of beneficial mutations. Over a period of 200 generations, the yeast populations displayed parallel evolutionary dynamics that are driven by the coexistence of independent beneficial mutations. Selective amplifications rapidly evolve under this selection pressure, in particular common inverted amplifications containing the sulfate transporter gene SUL1. Compared to single clones, detailed analysis of the populations uncovers a greater complexity whereby multiple subpopulations arise and compete despite a strong selection. The most common evolutionary adaptation to strong selection in these populations grown in sulfate limitation is determined by clonal interference, with adaptive variants both persisting and replacing one another.
Project description:Saccharomyces cerevisiae is an established microbial host for the production of non-native compounds. The synthesis of these compounds typically demands energy and competes with growth for carbon and energy substrate. Uncoupling product formation form growth would benefit product yields and decrease formation of by-product biomass. Studying non-growing metabolically-active yeast cultures provides a first step towards developing S. cerevisiae as a non-growing, robust cell factory. Non-growing metabolically-active cultures can be obtained in retentostat, a glucose-limited, continuous bioreactor system in which biomass accumulates while spent medium is constantly removed. Hitherto retentostat cultures of S. cerevisiae have only been reported under anaerobiosis, condition inappropriate for the production of energy-demanding products. The present study, using retentostat cultures, explores the physiology of non-dividing, fully respiring S. cerevisiae, focusing on industrially-relevant features. Following model-aided experimental design, retentostat cultivations were optimized for accelerated but smooth transition of S. cerevisiae from exponential growth to near-zero growth rates. During 20 days in retentostat the biomass concentration increased, leading very slow growth rates (specific growth rates below 0.001 h-1) but high culture viability (over 80% of viable cells). The maintenance requirement (mATP) was estimated at 0.64 mmolATP.gX-1.h-1, which is remarkably ca. 35% lower than the mATP measured in anaerobic retentostat cultures. Transcriptional down-regulation of genes involved in biosynthesis and up-regulation of stress-responsive genes towards near-zero growth rates corresponded well with data from anaerobic retentostats. More striking was the extreme heat-shock tolerance of S. cerevisiae, which exceeded by far previously reported heat shock tolerance of notoriously robust yeast cultures such as stationary phase cultures. Furthermore, while the metabolic fluxes in the retentostats were relatively low as a result of extreme caloric restriction, off-line measurements revealed that S. cerevisiae retained a high catabolic capacity. The high viability and extreme heat-shock tolerance revealed the robustness of S. cerevisiae at near-zero growth in retentostat. In addition, the relatively low maintenance requirements and high metabolic capacity under severe calorie restriction underline the potential of S. cerevisiae as a non-dividing microbial cell factory for the production of energy-intensive compounds. The retentostat is a promising tool to identify the molecular basis of this extreme robustness.
Project description:We examined the gene expression changes resulting from the evolution of resistance in experimental populations of the yeast Saccharomyces cerevisiae subjected to two antifungal drugs, fluconazole (FLC) and amphotericin B (AmB). Fluconazole resistance may involve increased efflux or changes in sterol metabolism, while AmB resistance generally involves changes in sterol metabolism; for all of these types of resistance, the gene expression changes are extensive. The goal of these experiments was to test whether failure of gene expression changes all downstream of the original mutation for drug resistance would affect the ability of a mutant cell to evolve and/or to support a drug-resistant phenotype.
Project description:Saccharomyces cerevisiae is an excellent microorganism for industrial succinic acid production, but high succinic acid concentration will inhibit the growth of Saccharomyces cerevisiae then reduce the production of succinic acid. Through analysis the transcriptomic data of Saccharomyces cerevisiae with different genetic backgrounds under different succinic acid stress, we hope to find the response mechanism of Saccharomyces cerevisiae to succinic acid.