In vivo continuous evolution of genes and pathways in yeast

Nathan Crook, Joseph Abatemarco, Jie Sun, James M. Wagner, Alexander Schmitz and Hal S. Alper ()
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Nathan Crook: The University of Texas at Austin
Joseph Abatemarco: The University of Texas at Austin
Jie Sun: The University of Texas at Austin
James M. Wagner: The University of Texas at Austin
Alexander Schmitz: The University of Texas at Austin
Hal S. Alper: The University of Texas at Austin

Nature Communications, 2016, vol. 7, issue 1, 1-14

Abstract: Abstract Directed evolution remains a powerful, highly generalizable approach for improving the performance of biological systems. However, implementations in eukaryotes rely either on in vitro diversity generation or limited mutational capacities. Here we synthetically optimize the retrotransposon Ty1 to enable in vivo generation of mutant libraries up to 1.6 × 107 l−1 per round, which is the highest of any in vivo mutational generation approach in yeast. We demonstrate this approach by using in vivo-generated libraries to evolve single enzymes, global transcriptional regulators and multi-gene pathways. When coupled to growth selection, this approach enables in vivo continuous evolution (ICE) of genes and pathways. Through a head-to-head comparison, we find that ICE libraries yield higher-performing variants faster than error-prone PCR-derived libraries. Finally, we demonstrate transferability of ICE to divergent yeasts, including Kluyveromyces lactis and alternative S. cerevisiae strains. Collectively, this work establishes a generic platform for rapid eukaryotic-directed evolution across an array of target cargo.

Date: 2016
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DOI: 10.1038/ncomms13051

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