Synthetic evolution

Article metrics

Abstract

The combination of modern biotechnologies such as DNA synthesis, λ red recombineering, CRISPR-based editing and next-generation high-throughput sequencing increasingly enables precise manipulation of genes and genomes. Beyond rational design, these technologies also enable the targeted, and potentially continuous, introduction of multiple mutations. While this might seem to be merely a return to natural selection, the ability to target evolution greatly reduces fitness burdens and focuses mutation and selection on those genes and traits that best contribute to a desired phenotype, ultimately throwing evolution into fast forward.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Fig. 1: Directed mutagenesis of targeted genes.
Fig. 2: Compartmentalization-mediated evolution.
Fig. 3: Shuffling-based hybridization of genes and genomes.
Fig. 4: Recombineering-based continuous evolution.
Fig. 5: CRISPR tools for genome evolution.
Fig. 6: Continuous evolution by host-mediated mutagenesis.
Fig. 7: Continuous self-evolution.

References

  1. 1.

    Thomason, L. C., Sawitzke, J. A., Li, X., Costantino, N. & Court, D. L. Recombineering: genetic engineering in bacteria using homologous recombination. Curr. Protoc. Mol. Biol. 106, 1.16.11–11.16.39 (2014).

  2. 2.

    Cong, L. et al. Multiplex genome engineering using CRISPR/Cas systems. Science 339, 819–823 (2013).

  3. 3.

    Doudna, J. A. & Charpentier, E. Genome editing. The new frontier of genome engineering with CRISPR-Cas9. Science 346, 1258096 (2014).

  4. 4.

    Jinek, M. et al. A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity. Science 337, 816–821 (2012).

  5. 5.

    Kumar, S. et al. Advanced selection methodologies for DNAzymes in sensing and healthcare applications. Trends Biochem. Sci. 44, 190–213 (2019).

  6. 6.

    Popović, M., Fliss, P. S. & Ditzler, M. A. In vitro evolution of distinct self-cleaving ribozymes in diverse environments. Nucleic Acids Res. 43, 7070–7082 (2015).

  7. 7.

    Aquino-Jarquin, G. & Toscano-Garibay, J. D. RNA aptamer evolution: two decades of SELEction. Int. J. Mol. Sci. 12, 9155–9171 (2011).

  8. 8.

    Kaur, H. Recent developments in cell-SELEX technology for aptamer selection. Biochim. Biophys. Acta Gen. Subj. 1862, 2323–2329 (2018).

  9. 9.

    Liu, R., Li, X. & Lam, K. S. Combinatorial chemistry in drug discovery. Curr. Opin. Chem. Biol. 38, 117–126 (2017).

  10. 10.

    Singh, H., Sharma, N., Khinchi, M., Gautam, S. & Kumawat, A. Combinatorial chemistry: a review. Asian J. Pharm. Res. Dev. (2017).

  11. 11.

    Dietrich, J. A., McKee, A. E. & Keasling, J. D. High-throughput metabolic engineering: advances in small-molecule screening and selection. Annu. Rev. Biochem. 79, 563–590 (2010).

  12. 12.

    Reidhaar-Olson, J. F. & Sauer, R. T. Combinatorial cassette mutagenesis as a probe of the informational content of protein sequences. Science 241, 53–57 (1988).

  13. 13.

    Oliphant, A. R. & Struhl, K. An efficient method for generating proteins with altered enzymatic properties: application to beta-lactamase. Proc. Natl Acad. Sci. USA 86, 9094–9098 (1989).

  14. 14.

    Palzkill, T. & Botstein, D. Identification of amino acid substitutions that alter the substrate specificity of TEM-1 beta-lactamase. J. Bacteriol. 174, 5237–5243 (1992).

  15. 15.

    Evnin, L. B., Vásquez, J. R. & Craik, C. S. Substrate specificity of trypsin investigated by using a genetic selection. Proc. Natl Acad. Sci. USA 87, 6659–6663 (1990).

  16. 16.

    Graham, L. D. et al. Random mutagenesis of the substrate-binding site of a serine protease can generate enzymes with increased activities and altered primary specificities. Biochemistry 32, 6250–6258 (1993).

  17. 17.

    Beckman, R. A., Mildvan, A. S. & Loeb, L. A. On the fidelity of DNA replication: manganese mutagenesis in vitro. Biochemistry 24, 5810–5817 (1985).

  18. 18.

    Cadwell, R. C. & Joyce, G. F. Randomization of genes by PCR mutagenesis. PCR Methods Appl. 2, 28–33 (1992).

  19. 19.

    Leung, D. W. A method for random mutagenesis of a defined DNA segment using a modified polymerase chain reaction. Technique 1, 11–15 (1989).

  20. 20.

    Zaccolo, M., Williams, D. M., Brown, D. M. & Gherardi, E. An approach to random mutagenesis of DNA using mixtures of triphosphate derivatives of nucleoside analogues. J. Mol. Biol. 255, 589–603 (1996).

  21. 21.

    Chen, K. Q. & Arnold, F. H. Enzyme engineering for nonaqueous solvents: random mutagenesis to enhance activity of subtilisin E in polar organic media. Bio/Technology 9, 1073–1077 (1991).

  22. 22.

    Bloom, J. D. et al. Evolving strategies for enzyme engineering. Curr. Opin. Struct. Biol. 15, 447–452 (2005).

  23. 23.

    Chen, W. & Struhl, K. Saturation mutagenesis of a yeast his3 “TATA element”: genetic evidence for a specific TATA-binding protein. Proc. Natl Acad. Sci. USA 85, 2691–2695 (1988).

  24. 24.

    Beaudry, A. A. & Joyce, G. F. Directed evolution of an RNA enzyme. Science 257, 635–641 (1992).

  25. 25.

    Service, R. F. Protein evolution earns chemistry Nobel. Science 362, 142 (2018).

  26. 26.

    Tawfik, D. S. & Griffiths, A. D. Man-made cell-like compartments for molecular evolution. Nat. Biotechnol. 16, 652–656 (1998).

  27. 27.

    Ghadessy, F. J., Ong, J. L. & Holliger, P. Directed evolution of polymerase function by compartmentalized self-replication. Proc. Natl Acad. Sci. USA 98, 4552–4557 (2001).

  28. 28.

    d’Abbadie, M. et al. Molecular breeding of polymerases for amplification of ancient DNA. Nat. Biotechnol. 25, 939–943 (2007).

  29. 29.

    Loakes, D., Gallego, J., Pinheiro, V. B., Kool, E. T. & Holliger, P. Evolving a polymerase for hydrophobic base analogues. J. Am. Chem. Soc. 131, 14827–14837 (2009).

  30. 30.

    Ellefson, J. W. et al. Synthetic evolutionary origin of a proofreading reverse transcriptase. Science 352, 1590–1593 (2016).

  31. 31.

    Williams, R. et al. Amplification of complex gene libraries by emulsion PCR. Nat. Methods 3, 545–550 (2006).

  32. 32.

    Ellefson, J. W. et al. Directed evolution of genetic parts and circuits by compartmentalized partnered replication. Nat. Biotechnol. 32, 97–101 (2014).

  33. 33.

    Meyer, A. J., Ellefson, J. W. & Ellington, A. D. Directed evolution of a panel of orthogonal T7 RNA polymerase variants for in vivo or in vitro synthetic circuitry. ACS Synth. Biol. 4, 1070–1076 (2015).

  34. 34.

    Ellefson, J. W., Ledbetter, M. P. & Ellington, A. D. Directed evolution of a synthetic phylogeny of programmable Trp repressors. Nat. Chem. Biol. 14, 361–367 (2018).

  35. 35.

    Meyer, A. J., Segall-Shapiro, T. H., Glassey, E., Zhang, J. & Voigt, C. A. Escherichia coli “Marionette” strains with 12 highly optimized small-molecule sensors. Nat. Chem. Biol. 15, 196–204 (2019).

  36. 36.

    Abil, Z., Ellefson, J. W., Gollihar, J. D., Watkins, E. & Ellington, A. D. Compartmentalized partnered replication for the directed evolution of genetic parts and circuits. Nat. Protoc. 12, 2493–2512 (2017).

  37. 37.

    Stemmer, W. P. DNA shuffling by random fragmentation and reassembly: in vitro recombination for molecular evolution. Proc. Natl Acad. Sci. USA 91, 10747–10751 (1994).

  38. 38.

    Stemmer, W. P. C. Rapid evolution of a protein in vitro by DNA shuffling. Nature 370, 389–391 (1994).

  39. 39.

    Crameri, A., Raillard, S.-A., Bermudez, E. & Stemmer, W. P. C. DNA shuffling of a family of genes from diverse species accelerates directed evolution. Nature 391, 288–291 (1998).

  40. 40.

    Nixon, A. E., Ostermeier, M. & Benkovic, S. J. Hybrid enzymes: manipulating enzyme design. Trends Biotechnol. 16, 258–264 (1998).

  41. 41.

    Ostermeier, M., Nixon, A. E., Shim, J. H. & Benkovic, S. J. Combinatorial protein engineering by incremental truncation. Proc. Natl Acad. Sci. USA 96, 3562–3567 (1999).

  42. 42.

    Ostermeier, M., Shim, J. H. & Benkovic, S. J. A combinatorial approach to hybrid enzymes independent of DNA homology. Nat. Biotechnol. 17, 1205–1209 (1999).

  43. 43.

    Hiraga, K. & Arnold, F. H. General method for sequence-independent site-directed chimeragenesis. J. Mol. Biol. 330, 287–296 (2003).

  44. 44.

    Kikuchi, M., Ohnishi, K. & Harayama, S. Novel family shuffling methods for the in vitro evolution of enzymes. Gene 236, 159–167 (1999).

  45. 45.

    Peisajovich, S. G., Rockah, L. & Tawfik, D. S. Evolution of new protein topologies through multistep gene rearrangements. Nat. Genet. 38, 168–174 (2006).

  46. 46.

    Meyer, A. J., Ellefson, J. W. & Ellington, A. D. Library generation by gene shuffling. Curr. Protoc. Mol. Biol. 105, 15.12 (2014).

  47. 47.

    Abil, Z. & Ellington, A. D. Compartmentalized self-replication for evolution of a DNA polymerase. Curr. Protoc. Chem. Biol. 10, 1–17 (2018).

  48. 48.

    Alper, H. & Stephanopoulos, G. Global transcription machinery engineering: a new approach for improving cellular phenotype. Metab. Eng. 9, 258–267 (2007).

  49. 49.

    Tan, F. et al. Using global transcription machinery engineering (gTME) to improve ethanol tolerance of Zymomonas mobilis. Microb. Cell Fact. 15, 4 (2016).

  50. 50.

    Alper, H., Moxley, J., Nevoigt, E., Fink, G. R. & Stephanopoulos, G. Engineering yeast transcription machinery for improved ethanol tolerance and production. Science 314, 1565–1568 (2006).

  51. 51.

    Park, K.-S. et al. Phenotypic alteration of eukaryotic cells using randomized libraries of artificial transcription factors. Nat. Biotechnol. 21, 1208–1214 (2003).

  52. 52.

    Auerbach, C. Chemical mutagenesis. Biol. Rev. Camb. Philos. Soc. 24, 355–391 (1949).

  53. 53.

    Muller, H. J. Artificial transmutation of the gene. Science 66, 84–87 (1927).

  54. 54.

    Ahloowalia, B. S., Maluszynski, M. & Nichterlein, K. Global impact of mutation-derived varieties. Euphytica 135, 187–204 (2004).

  55. 55.

    Gokhale, D. V., Puntambekar, U. S. & Deobagkar, D. N. Protoplast fusion: a tool for intergeneric gene transfer in bacteria. Biotechnol. Adv. 11, 199–217 (1993).

  56. 56.

    Peberdy, J. F. Developments in protoplast fusion in fungi. Microbiol. Sci. 4, 108–114 (1987).

  57. 57.

    Kao, K. N., Constabel, F., Michayluk, M. R. & Gamborg, O. L. Plant protoplast fusion and growth of intergeneric hybrid cells. Planta 120, 215–227 (1974).

  58. 58.

    Zhang, Y.-X. et al. Genome shuffling leads to rapid phenotypic improvement in bacteria. Nature 415, 644–646 (2002).

  59. 59.

    Quandt, E. M., Deatherage, D. E., Ellington, A. D., Georgiou, G. & Barrick, J. E. Recursive genomewide recombination and sequencing reveals a key refinement step in the evolution of a metabolic innovation in Escherichia coli. Proc. Natl Acad. Sci. USA 111, 2217–2222 (2014).

  60. 60.

    Sharan, S. K., Thomason, L. C., Kuznetsov, S. G. & Court, D. L. Recombineering: a homologous recombination-based method of genetic engineering. Nat. Protoc. 4, 206–223 (2009).

  61. 61.

    Ellis, H. M., Yu, D., DiTizio, T. & Court, D. L. High efficiency mutagenesis, repair, and engineering of chromosomal DNA using single-stranded oligonucleotides. Proc. Natl Acad. Sci. USA 98, 6742–6746 (2001).

  62. 62.

    Murphy, K. C. Use of bacteriophage lambda recombination functions to promote gene replacement in. Escherichia coli. J. Bacteriol. 180, 2063–2071 (1998).

  63. 63.

    Zhang, Y., Buchholz, F., Muyrers, J. P. P. & Stewart, A. F. A new logic for DNA engineering using recombination in Escherichia coli. Nat. Genet. 20, 123–128 (1998).

  64. 64.

    Wang, H. H. et al. Programming cells by multiplex genome engineering and accelerated evolution. Nature 460, 894–898 (2009).

  65. 65.

    Gallagher, R. R., Li, Z., Lewis, A. O. & Isaacs, F. J. Rapid editing and evolution of bacterial genomes using libraries of synthetic DNA. Nat. Protoc. 9, 2301–2316 (2014).

  66. 66.

    Raman, S., Rogers, J. K., Taylor, N. D. & Church, G. M. Evolution-guided optimization of biosynthetic pathways. Proc. Natl Acad. Sci. USA 111, 17803–17808 (2014).

  67. 67.

    Nyerges, Á. et al. Directed evolution of multiple genomic loci allows the prediction of antibiotic resistance. Proc. Natl Acad. Sci. USA 115, E5726–E5735 (2018).

  68. 68.

    Warner, J. R., Reeder, P. J., Karimpour-Fard, A., Woodruff, L. B. A. & Gill, R. T. Rapid profiling of a microbial genome using mixtures of barcoded oligonucleotides. Nat. Biotechnol. 28, 856–862 (2010).

  69. 69.

    Wang, H. H. et al. Genome-scale promoter engineering by coselection MAGE. Nat. Methods 9, 591–593 (2012).

  70. 70.

    Carr, P. A. et al. Enhanced multiplex genome engineering through co-operative oligonucleotide co-selection. Nucleic Acids Res. 40, e132 (2012).

  71. 71.

    Thompson, D. B. et al. The future of multiplexed eukaryotic genome engineering. ACS Chem. Biol. 13, 313–325 (2018).

  72. 72.

    Kow, Y. W., Bao, G., Reeves, J. W., Jinks-Robertson, S. & Crouse, G. F. Oligonucleotide transformation of yeast reveals mismatch repair complexes to be differentially active on DNA replication strands. Proc. Natl Acad. Sci. USA 104, 11352–11357 (2007).

  73. 73.

    Moerschell, R. P., Tsunasawa, S. & Sherman, F. Transformation of yeast with synthetic oligonucleotides. Proc. Natl Acad. Sci. USA 85, 524–528 (1988).

  74. 74.

    Storici, F., Durham, C. L., Gordenin, D. A. & Resnick, M. A. Chromosomal site-specific double-strand breaks are efficiently targeted for repair by oligonucleotides in yeast. Proc. Natl Acad. Sci. USA 100, 14994–14999 (2003).

  75. 75.

    DiCarlo, J. E. et al. Yeast oligo-mediated genome engineering (YOGE). ACS Synth. Biol. 2, 741–749 (2013).

  76. 76.

    Barbieri, E. M., Muir, P., Akhuetie-Oni, B. O., Yellman, C. M. & Isaacs, F. J. Precise editing at DNA replication forks enables multiplex genome engineering in eukaryotes. Cell 171, 1453–1467.e13 (2017).

  77. 77.

    Knott, G. J. & Doudna, J. A. CRISPR-Cas guides the future of genetic engineering. Science 361, 866–869 (2018).

  78. 78.

    Ran, F. A. et al. Genome engineering using the CRISPR-Cas9 system. Nat. Protoc. 8, 2281–2308 (2013).

  79. 79.

    Jiang, W., Bikard, D., Cox, D., Zhang, F. & Marraffini, L. A. RNA-guided editing of bacterial genomes using CRISPR-Cas systems. Nat. Biotechnol. 31, 233–239 (2013).

  80. 80.

    Ronda, C., Pedersen, L. E., Sommer, M. O. A. & Nielsen, A. T. CRMAGE: CRISPR optimized MAGE recombineering. Sci. Rep. 6, 19452 (2016).

  81. 81.

    Li, Y. et al. Metabolic engineering of Escherichia coli using CRISPR-Cas9 meditated genome editing. Metab. Eng. 31, 13–21 (2015).

  82. 82.

    Garst, A. D. et al. Genome-wide mapping of mutations at single-nucleotide resolution for protein, metabolic and genome engineering. Nat. Biotechnol. 35, 48–55 (2017).

  83. 83.

    Liu, R. et al. Iterative genome editing of Escherichia coli for 3-hydroxypropionic acid production. Metab. Eng. 47, 303–313 (2018).

  84. 84.

    DiCarlo, J. E. et al. Genome engineering in Saccharomyces cerevisiae using CRISPR-Cas systems. Nucleic Acids Res. 41, 4336–4343 (2013).

  85. 85.

    Jakočiūnas, T., Pedersen, L. E., Lis, A. V., Jensen, M. K. & Keasling, J. D. CasPER, a method for directed evolution in genomic contexts using mutagenesis and CRISPR/Cas9. Metab. Eng. 48, 288–296 (2018).

  86. 86.

    Roy, K. R. et al. Multiplexed precision genome editing with trackable genomic barcodes in yeast. Nat. Biotechnol. 36, 512–520 (2018).

  87. 87.

    Sadhu, M. J. et al. Highly parallel genome variant engineering with CRISPR-Cas9. Nat. Genet. 50, 510–514 (2018).

  88. 88.

    Han, K. et al. Synergistic drug combinations for cancer identified in a CRISPR screen for pairwise genetic interactions. Nat. Biotechnol. 35, 463–474 (2017).

  89. 89.

    Ma, L. et al. CRISPR-Cas9-mediated saturated mutagenesis screen predicts clinical drug resistance with improved accuracy. Proc. Natl Acad. Sci. USA 114, 11751–11756 (2017).

  90. 90.

    Findlay, G. M. et al. Accurate classification of BRCA1 variants with saturation genome editing. Nature 562, 217–222 (2018).

  91. 91.

    Komor, A. C., Kim, Y. B., Packer, M. S., Zuris, J. A. & Liu, D. R. Programmable editing of a target base in genomic DNA without double-stranded DNA cleavage. Nature 533, 420–424 (2016).

  92. 92.

    Gaudelli, N. M. et al. Programmable base editing of A•T to G•C in genomic DNA without DNA cleavage. Nature 551, 464–471 (2017).

  93. 93.

    Ma, Y. et al. Targeted AID-mediated mutagenesis (TAM) enables efficient genomic diversification in mammalian cells. Nat. Methods 13, 1029–1035 (2016).

  94. 94.

    Hess, G. T. et al. Directed evolution using dCas9-targeted somatic hypermutation in mammalian cells. Nat. Methods 13, 1036–1042 (2016).

  95. 95.

    Halperin, S. O. et al. CRISPR-guided DNA polymerases enable diversification of all nucleotides in a tunable window. Nature 560, 248–252 (2018).

  96. 96.

    Greener, A., Callahan, M. & Jerpseth, B. in In Vitro Mutagenesis Protocols (ed. Trower, M.K.) 375–385 (Humana, 1996).

  97. 97.

    Rasila, T. S., Pajunen, M. I. & Savilahti, H. Critical evaluation of random mutagenesis by error-prone polymerase chain reaction protocols, Escherichia coli mutator strain, and hydroxylamine treatment. Anal. Biochem. 388, 71–80 (2009).

  98. 98.

    Cumbers, S. J. et al. Generation and iterative affinity maturation of antibodies in vitro using hypermutating B-cell lines. Nat. Biotechnol. 20, 1129–1134 (2002).

  99. 99.

    Wang, L., Jackson, W. C., Steinbach, P. A. & Tsien, R. Y. Evolution of new nonantibody proteins via iterative somatic hypermutation. Proc. Natl Acad. Sci. USA 101, 16745–16749 (2004).

  100. 100.

    Camps, M., Naukkarinen, J., Johnson, B. P. & Loeb, L. A. Targeted gene evolution in Escherichia coli using a highly error-prone DNA polymerase I. Proc. Natl Acad. Sci. USA 100, 9727–9732 (2003).

  101. 101.

    Ravikumar, A., Arrieta, A. & Liu, C. C. An orthogonal DNA replication system in yeast. Nat. Chem. Biol. 10, 175–177 (2014).

  102. 102.

    Ravikumar, A., Arzumanyan, G. A., Obadi, M. K. A., Javanpour, A. A. & Liu, C. C. Scalable, continuous evolution of genes at mutation rates above genomic error thresholds. Cell 175, 1946–1957.e13 (2018).

  103. 103.

    Badran, A. H. & Liu, D. R. Development of potent in vivo mutagenesis plasmids with broad mutational spectra. Nat. Commun. 6, 8425 (2015).

  104. 104.

    Esvelt, K. M., Carlson, J. C. & Liu, D. R. A system for the continuous directed evolution of biomolecules. Nature 472, 499–503 (2011).

  105. 105.

    Carlson, J. C., Badran, A. H., Guggiana-Nilo, D. A. & Liu, D. R. Negative selection and stringency modulation in phage-assisted continuous evolution. Nat. Chem. Biol. 10, 216–222 (2014).

  106. 106.

    Packer, M. S., Rees, H. A. & Liu, D. R. Phage-assisted continuous evolution of proteases with altered substrate specificity. Nat. Commun. 8, 956 (2017).

  107. 107.

    Pu, J., Zinkus-Boltz, J. & Dickinson, B. C. Evolution of a split RNA polymerase as a versatile biosensor platform. Nat. Chem. Biol. 13, 432–438 (2017).

  108. 108.

    Bryson, D. I. et al. Continuous directed evolution of aminoacyl-tRNA synthetases. Nat. Chem. Biol. 13, 1253–1260 (2017).

  109. 109.

    Hu, J. H. et al. Evolved Cas9 variants with broad PAM compatibility and high DNA specificity. Nature 556, 57–63 (2018).

  110. 110.

    Badran, A. H. et al. Continuous evolution of Bacillus thuringiensis toxins overcomes insect resistance. Nature 533, 58–63 (2016).

  111. 111.

    Wang, T., Badran, A. H., Huang, T. P. & Liu, D. R. Continuous directed evolution of proteins with improved soluble expression. Nat. Chem. Biol. 14, 972–980 (2018).

  112. 112.

    Tschulena, U., Peterson, K. R., Gonzalez, B., Fedosyuk, H. & Barbas, C. F. III Positive selection of DNA-protein interactions in mammalian cells through phenotypic coupling with retrovirus production. Nat. Struct. Mol. Biol. 16, 1195–1199 (2009).

  113. 113.

    Berman, C. M. et al. An adaptable platform for directed evolution in human cells. J. Am. Chem. Soc. 140, 18093–18103 (2018).

  114. 114.

    Kalhor, R., Mali, P. & Church, G. M. Rapidly evolving homing CRISPR barcodes. Nat. Methods 14, 195–200 (2017).

  115. 115.

    Kalhor, R. et al. Developmental barcoding of whole mouse via homing CRISPR. Science 361, eaat9804 (2018).

  116. 116.

    van Opijnen, T., Bodi, K. L. & Camilli, A. Tn-seq: high-throughput parallel sequencing for fitness and genetic interaction studies in microorganisms. Nat. Methods 6, 767–772 (2009).

  117. 117.

    Crook, N. et al. In vivo continuous evolution of genes and pathways in yeast. Nat. Commun. 7, 13051 (2016).

  118. 118.

    Gabriel, A., Willems, M., Mules, E. H. & Boeke, J. D. Replication infidelity during a single cycle of Ty1 retrotransposition. Proc. Natl Acad. Sci. USA 93, 7767–7771 (1996).

  119. 119.

    Boutabout, M., Wilhelm, M. & Wilhelm, F. X. DNA synthesis fidelity by the reverse transcriptase of the yeast retrotransposon Ty1. Nucleic Acids Res. 29, 2217–2222 (2001).

  120. 120.

    Lampson, B. C., Inouye, M. & Inouye, S. Retrons, msDNA, and the bacterial genome. Cytogenet. Genome Res. 110, 491–499 (2005).

  121. 121.

    Farzadfard, F. & Lu, T. K. Genomically encoded analog memory with precise in vivo DNA writing in living cell populations. Science 346, 1256272 (2014).

  122. 122.

    Simon, A. J., Morrow, B. R. & Ellington, A. D. Retroelement-based genome editing and evolution. ACS Synth. Biol. 7, 2600–2611 (2018).

  123. 123.

    Zetsche, B. et al. Multiplex gene editing by CRISPR-Cpf1 using a single crRNA array. Nat. Biotechnol. 35, 31–34 (2017).

  124. 124.

    Hupfeld, M. et al. A functional type II-A CRISPR-Cas system from Listeria enables efficient genome editing of large non-integrating bacteriophage. Nucleic Acids Res. 46, 6920–6933 (2018).

  125. 125.

    Wright, A. V., Nuñez, J. K. & Doudna, J. A. Biology and applications of CRISPR systems: harnessing nature’s toolbox for genome engineering. Cell 164, 29–44 (2016).

  126. 126.

    Yan, W. X. et al. Functionally diverse type V CRISPR-Cas systems. Science 363, 88–91 (2019).

  127. 127.

    Wu, L. et al. Diversity-generating retroelements: natural variation, classification and evolution inferred from a large-scale genomic survey. Nucleic Acids Res. 46, 11–24 (2018).

  128. 128.

    Zimmerly, S. & Wu, L. An unexplored diversity of reverse transcriptases in bacteria. Microbiol. Spectr. 3, MDNA3-0058-2014 (2015).

  129. 129.

    Liu, M. et al. Reverse transcriptase-mediated tropism switching in Bordetella bacteriophage. Science 295, 2091–2094 (2002).

  130. 130.

    Medhekar, B. & Miller, J. F. Diversity-generating retroelements. Curr. Opin. Microbiol. 10, 388–395 (2007).

  131. 131.

    Doulatov, S. et al. Tropism switching in Bordetella bacteriophage defines a family of diversity-generating retroelements. Nature 431, 476–481 (2004).

  132. 132.

    Paul, B. G. et al. Targeted diversity generation by intraterrestrial archaea and archaeal viruses. Nat. Commun. 6, 6585 (2015).

  133. 133.

    Paul, B. G. et al. Retroelement-guided protein diversification abounds in vast lineages of Bacteria and Archaea. Nat. Microbiol. 2, 17045 (2017).

  134. 134.

    Yuan, T. Z., Overstreet, C. M., Moody, I. S. & Weiss, G. A. Protein engineering with biosynthesized libraries from Bordetella bronchiseptica bacteriophage. PLoS One 8, e55617 (2013).

  135. 135.

    Dymond, J. S. et al. Synthetic chromosome arms function in yeast and generate phenotypic diversity by design. Nature 477, 471–476 (2011).

  136. 136.

    Jia, B. et al. Precise control of SCRaMbLE in synthetic haploid and diploid yeast. Nat. Commun. 9, 1933 (2018).

  137. 137.

    Enyeart, P. J. et al. Generalized bacterial genome editing using mobile group II introns and Cre-lox. Mol. Syst. Biol. 9, 685 (2013).

  138. 138.

    Gao, L. et al. Engineered Cpf1 variants with altered PAM specificities. Nat. Biotechnol. 35, 789–792 (2017).

  139. 139.

    Mason, D. M. et al. High-throughput antibody engineering in mammalian cells by CRISPR/Cas9-mediated homology-directed mutagenesis. Nucleic Acids Res. 46, 7436–7449 (2018).

  140. 140.

    Akcakaya, P. et al. In vivo CRISPR editing with no detectable genome-wide off-target mutations. Nature 561, 416–419 (2018).

Download references

Acknowledgements

This work was funded by the Air Force Office of Scientific Research (FA9550-14-1-0089) and by an Arnold O. Beckman Postdoctoral Fellowship held by A.J.S.

Author information

Correspondence to Andrew D. Ellington.

Ethics declarations

Competing interests

A.D.E is cofounder and advisor at GRO Biosciences, Inc, which is using computational protein design and synthetic biology to develop protein therapeutics. A.D.E has filed intellectual property disclosures that reference Compartmentalized Partner Replication: 6761ELL ‘Thermostable reverse transcriptase based on a thermostable DNA polymerase,’ US 15/410, 211, Japan 2018-538718 and EP 17741900.9, filed on 1/19/17; and 7151 ELL ‘A method for screening metabolites and their receptors’ PCT/US2018/037818, filed on 6/15/18.

Additional information

Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Simon, A.J., d’Oelsnitz, S. & Ellington, A.D. Synthetic evolution. Nat Biotechnol 37, 730–743 (2019) doi:10.1038/s41587-019-0157-4

Download citation

Further reading