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Updated: Jul 4, 2026
Growth-based Determination and Biochemical Confirmation of Genetic Requirements for Protein Degradation in Saccharomyces cerevisiae
Published on: February 16, 2015
Plasmid instability kinetics of the yeast S288C pUCKm8 [cir+] in non-selective and selective media
L S Schwartz1, N B Jansen, N W Ho
1Laboratory of Renewable Resources Engineering, Purdue University, West Lafayette, Indiana 47907, USA.
Abstract:
Plasmid loss kinetics for Saccharomyces cerevisiae transformed with the 2-mum DNA-based-plasmid pUCKm8 were measured in nonselective and selective media. The plasmid pUCKm8 gives the organism two new phenotypes: resistance to the wide spectrum antibiotic G418 sulfate, and the ability to produce the enzyme, beta-lactamase. Plasmid stability was determined using the production of beta-lactamase as a marker. The effect of G418 on the growth rates of all organisms present in the culture and on plasmid stability was also determined. Mathematical models describing plasmid loss kinetics during exponential growth for both nonselective and selective conditions are used to simulate the experimental data. In nonselective medium, over 80% of the cells still exhibited the desired phenotype after 50 doublings. In medium containing G418, improvements in plasmid stability were only marginal due to the appearance of antibiotic-resistant cells.
Insights
This study investigated plasmid loss in yeast, finding that the pUCKm8 plasmid remained stable in over 80% of cells after 50 doublings in nonselective media. However, G418 antibiotic only marginally improved plasmid stability due to resistant cells.
Area of Science:
- * Molecular Biology
- * Microbiology
- * Yeast Genetics
Background:
- * Saccharomyces cerevisiae is a widely used model organism in genetic research.
- * Plasmids are extrachromosomal DNA elements crucial for introducing new traits into host cells.
- * The 2-micron DNA-based plasmid pUCKm8 confers antibiotic resistance and beta-lactamase production in yeast.
Purpose of the Study:
- * To quantify plasmid loss kinetics of pUCKm8 in Saccharomyces cerevisiae.
- * To evaluate the impact of selective (G418 sulfate) and nonselective media on plasmid stability.
- * To assess the effect of G418 sulfate on yeast growth rates and plasmid stability.
Main Methods:
- * Measurement of plasmid loss kinetics in Saccharomyces cerevisiae transformed with pUCKm8.
- * Use of beta-lactamase production as a marker for plasmid stability.
- * Application of mathematical models to simulate plasmid loss during exponential growth.
- * Determination of G418 sulfate's effect on growth rates and plasmid stability.
Main Results:
- * In nonselective medium, over 80% of yeast cells retained the pUCKm8 plasmid after 50 doublings.
- * Plasmid stability showed only marginal improvement in G418 sulfate-containing medium.
- * The presence of antibiotic-resistant cells in selective media complicated the assessment of plasmid stability.
Conclusions:
- * The pUCKm8 plasmid exhibits high stability in Saccharomyces cerevisiae under nonselective conditions.
- * G418 sulfate offers limited enhancement of plasmid stability due to the emergence of resistant yeast strains.
- * Mathematical modeling provides a robust framework for analyzing plasmid loss kinetics in microbial systems.
