Yeast oligo-mediated genome engineering (YOGE)

James E. Dicarlo, Andrew J. Conley, Merja Penttilä, Jussi Jäntti, Harris H. Wang, George M. Church (Corresponding Author)

    Research output: Contribution to journalArticleScientificpeer-review

    102 Citations (Scopus)

    Abstract

    High-frequency oligonucleotide-directed recombination engineering (recombineering) has enabled rapid modification of several prokaryotic genomes to date. Here, we present a method for oligonucleotide-mediated recombineering in the model eukaryote and industrial production host Saccharomyces cerevisiae, which we call yeast oligo-mediated genome engineering (YOGE). Through a combination of overexpression and knockouts of relevant genes and optimization of transformation and oligonucleotide designs, we achieve high gene-modification frequencies at levels that only require screening of dozens of cells. We demonstrate the robustness of our approach in three divergent yeast strains, including those involved in industrial production of biobased chemicals. Furthermore, YOGE can be iteratively executed via cycling to generate genomic libraries up to 105 individuals at each round for diversity generation. YOGE cycling alone or in combination with phenotypic selections or endonuclease-based negative genotypic selections can be used to generate modified alleles easily in yeast populations with high frequencies.

    Original languageEnglish
    Pages (from-to)741-749
    JournalACS Synthetic Biology
    Volume2
    Issue number12
    DOIs
    Publication statusPublished - 20 Dec 2013
    MoE publication typeA1 Journal article-refereed

    Keywords

    • genome engineering
    • MAGE
    • oligonucleotide transformation
    • recombineering
    • Saccharomyces cerevisiae
    • yeast

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