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The developing toolkit of continuous directed evolution

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Abstract

Continuous directed evolution methods allow the key steps of evolution—gene diversification, selection, and replication—to proceed in the laboratory with minimal researcher intervention. As a result, continuous evolution can find solutions much more quickly than traditional discrete evolution methods. Continuous evolution also enables the exploration of longer and more numerous evolutionary trajectories, increasing the likelihood of accessing solutions that require many steps through sequence space and greatly facilitating the iterative refinement of selection conditions and targeted mutagenesis strategies. Here we review the historical advances that have expanded continuous evolution from its earliest days as an experimental curiosity to its present state as a powerful and surprisingly general strategy for generating tailor-made biomolecules, and discuss more recent improvements with an eye to the future.

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Fig. 1: An overview of laboratory directed evolution, including major methods for each key step.
Fig. 2: Overview of early in vitro continuous evolution systems.
Fig. 3: Systems for continuous culture.
Fig. 4: Continuous evolution via orthogonal polymerases and reverse transcriptases.
Fig. 5: Phage-assisted continuous evolution (PACE).
Fig. 6: PACE selection strategies.
Fig. 7: Viral evolution of genetically actuating sequences (VEGAS).

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Acknowledgements

D.R.L. gratefully acknowledges support from NIH U01 AI142756 (D.R.L.), RM1 HG009490 (D.R.L.), R01 EB022376 (D.R.L.), and R35 GM118062 (D.R.L.); and HHMI (D.R.L.). We thank A. Badran and K. Zhao for their helpful comments.

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Authors and Affiliations

  1. Merkin Institute of Transformative Technologies in Healthcare, Broad Institute of MIT and Harvard, Cambridge, MA, USA

    Mary S. Morrison, Christopher J. Podracky & David R. Liu

  2. Howard Hughes Medical Institute, Harvard University, Cambridge, MA, USA

    Mary S. Morrison, Christopher J. Podracky & David R. Liu

  3. Department of Molecular and Cellular Biology, Harvard University, Cambridge, MA, USA

    Mary S. Morrison

  4. Department of Chemistry and Chemical Biology, Harvard University, Cambridge, MA, USA

    Mary S. Morrison, Christopher J. Podracky & David R. Liu

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  1. Mary S. Morrison
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Correspondence to David R. Liu.

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D.R.L. is a consultant and co-founder of Prime Medicine, Beam Therapeutics, Pairwise Plants, and Editas Medicine, companies that use genome editing. Complete disclosures are available at https://liugroup.us.

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Morrison, M.S., Podracky, C.J. & Liu, D.R. The developing toolkit of continuous directed evolution. Nat Chem Biol 16, 610–619 (2020). https://doi.org/10.1038/s41589-020-0532-y

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