Abstract
Rapid and highly efficient mating-type switching of Saccharomyces cerevisiae enables a wide variety of genetic manipulations, such as the construction of strains, for instance, isogenic haploid pairs of both mating-types, diploids and polyploids. We used the CRISPR/Cas9 system to generate a double-strand break at the MAT locus and, in a single cotransformation, both haploid and diploid cells were switched to the specified mating-type at ~80% efficiency. The mating-type of strains carrying either rod or ring chromosome III were switched, including those lacking HMLa and HMRa cryptic mating loci. Furthermore, we transplanted the synthetic yeast chromosome V to build a haploid polysynthetic chromosome strain by using this method together with an endoreduplication intercross strategy. The CRISPR/Cas9 mating-type switching method will be useful in building the complete synthetic yeast (Sc2.0) genome. Importantly, it is a generally useful method to build polyploids of a defined genotype and generally expedites strain construction, for example, in the construction of fully a/a/α/α isogenic tetraploids.
Original language | English (US) |
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Pages (from-to) | 173-183 |
Number of pages | 11 |
Journal | G3: Genes, Genomes, Genetics |
Volume | 8 |
Issue number | 1 |
DOIs | |
State | Published - Jan 1 2018 |
Keywords
- CRISPR/Cas9
- Mating-type switching
- Polyploidy
- Ring chromosome
- Saccharomyces cerevisiae
ASJC Scopus subject areas
- Molecular Biology
- Genetics
- Genetics(clinical)