The DOI matches its Crossref record; 2 of the 2 references it cites were found in a registry or an index.
A system for the continuous directed evolution of biomolecules
Open accessAbstract
Laboratory evolution has generated many biomolecules with desired properties, but a single round of mutation, gene expression, screening or selection, and replication typically requires days or longer with frequent human intervention. Because evolutionary success is dependent on the total number of rounds performed, a means of performing laboratory evolution continuously and rapidly could dramatically enhance its effectiveness. Although researchers have accelerated individual steps in the evolutionary cycle, the only previous example of continuous directed evolution was the landmark study of Wright and Joyce, who continuously evolved RNA ligase ribozymes with an in vitro replication cycle that unfortunately cannot be easily adapted to other biomolecules. Here we describe a system that enables the continuous directed evolution of gene-encoded molecules that can be linked to protein production in Escherichia coli. During phage-assisted continuous evolution (PACE), evolving genes are transferred from host cell to host cell through a modified bacteriophage life cycle in a manner that is dependent on the activity of interest. Dozens of rounds of evolution can occur in a single day of PACE without human intervention. Using PACE, we evolved T7 RNA polymerase (RNAP) variants that recognize a distinct promoter, initiate transcripts with ATP instead of GTP, and initiate transcripts with CTP. In one example, PACE executed 200 rounds of protein evolution over the course of 8 days. Starting from undetectable activity levels in two of these cases, enzymes with each of the three target activities emerged in less than 1 week of PACE. In all three cases, PACE-evolved polymerase activities exceeded or were comparable to that of the wild-type T7 RNAP on its wild-type promoter, representing improvements of up to several hundred-fold. By greatly accelerating laboratory evolution, PACE may provide solutions to otherwise intractable directed evolution problems and address novel questions about molecular evolution.
Overview
The claims and statistics of this paper haven’t been checked yet.
Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.
Integrity signals
Registry
Cites
Statistics
Verification
Claims the paper makes
The claims and statistics of this paper haven’t been checked yet.
Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.
Self-check
Percentages, sums and reported differences re-computed from the paper’s own numbers.
The self-check needs the paper’s full text.
Citations
Cites
Programming cells by multiplex genome engineering and accelerated evolution
Wang, H. H., Isaacs, F. J., Carr, P. A., Sun, Z. Z., Xu, G., Forest, C. R., & Church, G. M. (2009) · Nature · doi:10.1038/nature08187 · Cited by 1,529
An extracellular Darwinian experiment with a self-duplicating nucleic acid molecule.
Mills, D. R., Peterson, R. L., & Spiegelman, S. (1967) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.58.1.217 · Cited by 648
Cited by
RNA-Guided Human Genome Engineering via Cas9
Mali, P., Yang, L., Esvelt, K. M., Aach, J., Guell, M., DiCarlo, J. E., Norville, J. E., & Church, G. M. (2013) · Science · doi:10.1126/science.1232033 · Cited by 9,210
Self-citationProgrammable base editing of A•T to G•C in genomic DNA without DNA cleavage
Gaudelli, N. M., Komor, A. C., Rees, H. A., Packer, M. S., Badran, A. H., Bryson, D. I., & Liu, D. R. (2017) · Nature · doi:10.1038/nature24644 · Cited by 4,184
Self-citationEvolved Cas9 variants with broad PAM compatibility and high DNA specificity
Hu, J. H., Miller, S. M., Geurts, M. H., Tang, W., Chen, L., Sun, N., Zeina, C. M., Gao, X., Rees, H. A., Lin, Z., et al. (2018) · Nature · doi:10.1038/nature26155 · Cited by 1,552
Self-citationEnd-to-end design of wearable sensors
Ates, H. C., Nguyen, P. Q., Gonzalez-Macia, L., Morales-Narváez, E., Güder, F., Collins, J. J., & Dincer, C. (2022) · Nature Reviews Materials · doi:10.1038/s41578-022-00460-x · Cited by 1,306
Challenges and opportunities for structural DNA nanotechnology
Pinheiro, A. V., Han, D., Shih, W. M., & Yan, H. (2011) · Nature Nanotechnology · doi:10.1038/nnano.2011.187 · Cited by 1,292
Cas9 as a versatile tool for engineering biology
Mali, P., Esvelt, K. M., & Church, G. M. (2013) · Nature Methods · doi:10.1038/nmeth.2649 · Cited by 1,197
Self-citationRichter, M. F., Zhao, K. T., Eton, E., Lapinaite, A., Newby, G. A., Thuronyi, B. W., Wilson, C., Koblan, L. W., Zeng, J., Bauer, D. E., et al. (2020) · Nature Biotechnology · doi:10.1038/s41587-020-0453-z · Cited by 1,020
Self-citationPrebiotic Systems Chemistry: New Perspectives for the Origins of Life
Ruiz-Mirazo, K., Briones, C., & de la Escosura, A. (2014) · Chemical Reviews · doi:10.1021/cr2004844 · Cited by 930
Latest Developed Strategies to Minimize the Off-Target Effects in CRISPR-Cas-Mediated Genome Editing
Naeem, M., Majeed, S., Hoque, M. Z., & Ahmad, I. (2020) · Cells · doi:10.3390/cells9071608 · Cited by 457
Continuous evolution of base editors with expanded target compatibility and improved activity
Thuronyi, B. W., Koblan, L. W., Levy, J. M., Yeh, W.-H., Zheng, C., Newby, G. A., Wilson, C., Bhaumik, M., Shubina-Oleinik, O., Holt, J. R., et al. (2019) · Nature Biotechnology · doi:10.1038/s41587-019-0193-0 · Cited by 347
Self-citationGenomically encoded analog memory with precise in vivo DNA writing in living cell populations
Farzadfard, F., & Lu, T. K. (2014) · Science · doi:10.1126/science.1256272 · Cited by 298
Fourth generation biofuel from genetically modified algal biomass: Challenges and future directions.
Shokravi, H., Shokravi, Z., Heidarrezaei, M., Ong, H. C., Rahimian Koloor, S. S., Petrů, M., Lau, W. J., & Ismail, A. F. (2021) · Chemosphere · doi:10.1016/j.chemosphere.2021.131535 · Cited by 127
CRISPR engineering in organoids for gene repair and disease modelling
Geurts, M. H., & Clevers, H. (2023) · Nature Reviews Bioengineering · doi:10.1038/s44222-022-00013-5 · Cited by 103
Kunisch, F., Campobasso, C., Wagemans, J., Yildirim, S., Chan, B. K., Schaudinn, C., Lavigne, R., Turner, P. E., Raschke, M. J., Trampuz, A., et al. (2024) · Nature Communications · doi:10.1038/s41467-024-52595-w · Cited by 101
Light and carbon: Synthetic biology toward new cyanobacteria-based living biomaterials
Goodchild-Michelman, I. M., Church, G. M., Schubert, M. G., & Tang, T.-C. (2023) · Materials Today Bio · doi:10.1016/j.mtbio.2023.100583 · Cited by 41
Directed evolution of hyperactive integrases for site specific insertion of transgenes
Hew, B. E., Gupta, S., Sato, R., Waller, D. F., Stoytchev, I., Short, J. E., Sharek, L., Tran, C. T., Badran, A. H., & Owens, J. B. (2024) · Nucleic Acids Research · doi:10.1093/nar/gkae534 · Cited by 35
Integrating Computational Design and Experimental Approaches for Next-Generation Biologics
Son, A., Park, J., Kim, W., Lee, W., Yoon, Y., Ji, J., & Kim, H. (2024) · Biomolecules · doi:10.3390/biom14091073 · Cited by 29
Selecting Better Biocatalysts by Complementing Recoded Bacteria**
Rubini, R., Jansen, S. C., Beekhuis, H., Rozeboom, H. J., & Mayer, C. (2023) · Angewandte Chemie International Edition · doi:10.1002/anie.202213942 · Cited by 15
A chimeric viral platform for directed evolution in mammalian cells
Cole, A. J., Denes, C. E., Moreno, C. L., Hunault, L., Dobson, T., Hesselson, D., & Neely, G. G. (2025) · Nature Communications · doi:10.1038/s41467-025-59438-2 · Cited by 7
Deng, Y., Etheridge, K., Ran, X., Maurais, H. E., & Sarpeshkar, R. (2025) · Applied and Environmental Microbiology · doi:10.1128/aem.02491-24 · Cited by 7
Metadata
- Authors
- Esvelt, K. M., Carlson, J. C., & Liu, D. R.
- Venue
- Nature
- Published
- 2011-04-10
- Volume
- 472
- Issue
- 7344
- Pages
- 499-503
- ISSN
- 0028-0836 (ISSN-L)
- 1476-4687
- OpenAlex
- W2097861808
- Cited by
- 699
- References
- 2
- FWCI
- 18.27
- Type
- journal article
- Language
- en
- Open access
- green
About these checks
Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.
Statistics and arithmetic are recomputed, not judged.
Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.
What counts as a claim ›How we measure confidence ›How we check this ›
Are you an author, or do you see a problem with this page? Request a review
Data sources
Bibliographic data: Semantic Scholar, OpenAlex, Crossref.
Contains information from the Semantic Scholar Academic Graph Datasets, made available under the ODC Attribution License (ODC-BY 1.0).
Semantic Scholar’s data is described in: Kinney, R., et al. (2023). The Semantic Scholar Open Data Platform. arXiv:2301.10140.
