Rapid and efficient CRISPR/Cas9-based mating-type switching of Saccharomyces cerevisiae

Ze Xiong Xie, Leslie A. Mitchell, Hui Min Liu, Bing Zhi Li, Duo Liu, Neta Agmon, Yi Wu, Xia Li, Xiao Zhou, Bo Li, Wen Hai Xiao, Ming Zhu Ding, Ying Wang, Ying Jin Yuan, Jef D. Boeke

    Research output: Contribution to journalArticle

    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 languageEnglish (US)
    Pages (from-to)173-183
    Number of pages11
    JournalG3: Genes, Genomes, Genetics
    Volume8
    Issue number1
    DOIs
    StatePublished - Jan 1 2018

    Keywords

    • CRISPR/Cas9
    • Mating-type switching
    • Polyploidy
    • Ring chromosome
    • Saccharomyces cerevisiae

    ASJC Scopus subject areas

    • Molecular Biology
    • Genetics
    • Genetics(clinical)

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