Related Experiment Videos
Cycloheximide resistance as a yeast cloning marker.
L del Pozo1, D Abarca, M G Claros
1Centro de Biología Molecular (C.S.I.C.-U.A.M.), Universidad Autónoma de Madrid, Canto Blanco, Spain.
Current Genetics
|May 1, 1991
Summary
Two mutations in Saccharomyces cerevisiae confer cycloheximide (cyh) resistance. The study identified two genes, cyh2 and a new gene cyh5, responsible for high and low resistance, respectively, in yeast strains.
Area of Science:
- Genetics
- Molecular Biology
- Microbiology
Background:
- Cycloheximide (cyh) resistance in yeast Saccharomyces cerevisiae is often dominant in heterozygous diploids.
- A specific haploid strain, MMY1, confers high cyh-resistance to heterozygous diploids.
Purpose of the Study:
- To genetically and biochemically characterize the cyh-resistant yeast strain MMY1.
- To identify and clone the genes responsible for cycloheximide resistance in MMY1.
Main Methods:
- Tetrad analysis to study inheritance of resistance mutations.
- Genomic library construction and plasmid isolation.
- Pulse field gel electrophoresis and nucleotide sequencing for gene mapping and identification.
Main Results:
- MMY1 harbors two unlinked nuclear mutations conferring high and low cyh-resistance.
- Two plasmids, pRC1 and pRC13, were isolated, conferring high and low resistance, respectively.
- pRC1 contains the known cyh2 gene on chromosome VII; pRC13 contains a novel gene, cyh5, on chromosome XI.
Conclusions:
- The dominant cyh-resistance phenotype in MMY1 is due to the combined action of cyh2 and cyh5.
- Identified a previously unreported locus, cyh5, involved in cycloheximide resistance.
- Successfully cloned genes conferring distinct levels of cycloheximide resistance in yeast.