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  • Review Article
  • Published:

Genome synthesis in plants

Abstract

Owing to advances in genome sequencing and editing, a genome can now be redesigned, synthesized and introduced into cells as desired. The field of synthetic genomics not only aims to provide deeper understanding of how the genome functions but can also be harnessed for a wide range of synthetic biology and bioengineering applications, from rapid evolution and screening for favourable strains to biotechnological and bioproduction tool development. Although genome synthesis has been carried out mainly in simple unicellular organisms, plants and animals are now also being investigated. Compared with animals, plants have unique advantages, such as fewer ethical concerns, simpler experimental operations and easier regeneration from cells to organisms. In this Review, we focus on genome synthesis in plants, discuss the current research landscape and assess possible future directions.

Key points

  • Advances in genome sequencing, synthesis and editing technologies have enabled systematic redesign and chemical synthesis of plant genomes, establishing a new research discipline for functional genome exploration with potential for agricultural innovation and industrial bioproduction.

  • Top-down chromosomal engineering strategies focus on the modification of endogenous chromosomes, and bottom-up strategies use de novo DNA assembly techniques to generate artificial chromosomes with distinct structural and functional properties.

  • De novo synthesis of plant chromosome segments follows a stage-gated workflow involving computational design, modular DNA assembly, transformation, targeted integration and verification, plant regeneration and phenotypic validating, with post-analysis data driving iterative optimization cycles for genome design refinement.

  • Accurate integration of large DNA fragments remains a technical bottleneck in seed plant synthetic genomics, requiring breakthroughs in homology-directed repair or nonhomologous end joining-mediated integration efficiency and engineered recombinase systems to improve plant genome engineering platforms.

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Fig. 1: Timeline of milestones in synthetic genomics.
Fig. 2: Strategies of chromosome synthesis for multicellular organisms.
Fig. 3: Designing synthetic plant genomes.
Fig. 4: Artificial plant centromeres.
Fig. 5: Targeted replacement of large DNA fragments.

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References

  1. McKusick, V. A. & Ruddle, F. H. Toward a complete map of the human genome. Genomics 1, 103–106 (1987).

    Article  Google Scholar 

  2. Sinyakov, A. N., Ryabinin, V. A. & Kostina, E. V. Application of array-based oligonucleotides for synthesis of genetic designs. Mol. Biol. 55, 487–500 (2021).

    Article  Google Scholar 

  3. Casini, A., Storch, M., Baldwin, G. S. & Ellis, T. Bricks and blueprints: methods and standards for DNA assembly. Nat. Rev. Mol. Cell Biol. 16, 568–576 (2015).

    Article  Google Scholar 

  4. He, B. et al. YLC-assembly: large DNA assembly via yeast life cycle. Nucleic Acids Res. 51, 8283–8292 (2023).

    Article  Google Scholar 

  5. Cello, J., Paul, A. V. & Wimmer, E. Chemical synthesis of poliovirus cDNA: generation of infectious virus in the absence of natural template. Science 297, 1016–1018 (2002). This study established the first fully synthetic genome, validating template-independent de novo genome construction through chemical synthesis methodologies.

    Article  Google Scholar 

  6. Smith, H. O., Hutchison, C. A., Pfannkoch, C. & Venter, J. C. Generating a synthetic genome by whole genome assembly: φX174 bacteriophage from synthetic oligonucleotides. Proc. Natl Acad. Sci. USA 100, 15440–15445 (2003).

    Article  Google Scholar 

  7. Gibson, D. G. et al. Creation of a bacterial cell controlled by a chemically synthesized genome. Science 329, 52–56 (2010).

    Article  Google Scholar 

  8. Fredens, J. et al. Total synthesis of Escherichia coli with a recoded genome. Nature 569, 514–518 (2019).

    Article  Google Scholar 

  9. Hutchison, C. A. et al. Design and synthesis of a minimal bacterial genome. Science 351, aad6253 (2016). This study reports the excision of dispensable genes (47% reduction from 901 to 473) through systematic genome streamlining, resulting in the first functional minimal synthetic cell prototype.

    Article  Google Scholar 

  10. Zhang, W., Mitchell, L. A., Bader, J. S. & Boeke, J. D. Synthetic genomes. Annu. Rev. Biochem. 89, 77–101 (2020).

    Article  Google Scholar 

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

    Article  Google Scholar 

  12. Annaluru, N. et al. Total synthesis of a functional designer eukaryotic chromosome. Science 344, 55–58 (2014).

    Article  Google Scholar 

  13. Richardson, S. M. et al. Design of a synthetic yeast genome. Science 355, 1040–1044 (2017). This study describes the Sc2.0 project, which delivered a definitive blueprint for synthetic eukaryotic genomics, incorporating sequence-editing strategies, synthetic telomere design and inducible evolution systems.

    Article  Google Scholar 

  14. Zhang, W. et al. Mouse genome rewriting and tailoring of three important disease loci. Nature 623, 423–431 (2023).

    Article  Google Scholar 

  15. Chen, L.-G. et al. A designer synthetic chromosome fragment functions in moss. Nat. Plants 10, 228–239 (2024). This study is the first demonstration of partial chromosome arm synthesis in plants.

    Article  Google Scholar 

  16. Boeke, J. D. et al. The genome project-write. Science 353, 126–127 (2016).

    Article  Google Scholar 

  17. Jiao, Y. & Wang, Y. Towards plant synthetic genomics. Biodes. Res. 5, 0020 (2023).

    Article  Google Scholar 

  18. Blommaert, J. Genome size evolution: towards new model systems for old questions. Proc. Biol. Sci. 287, 20201441 (2020).

    Google Scholar 

  19. AGARWAL, K. L. et al. Total synthesis of the gene for an alanine transfer ribonucleic acid from yeast. Nature 227, 27–34 (1970).

    Article  Google Scholar 

  20. Koonin, E. V., Dolja, V. V. & Krupovic, M. The logic of virus evolution. Cell Host Microbe 30, 917–929 (2022).

    Article  Google Scholar 

  21. Racaniello, V. R. & Baltimore, D. Molecular cloning of poliovirus cDNA and determination of the complete nucleotide sequence of the viral genome. Proc. Natl Acad. Sci. USA 78, 4887–4891 (1981).

    Article  Google Scholar 

  22. Venter, J. C., Glass, J. I., Hutchison, C. A. & Vashee, S. Synthetic chromosomes, genomes, viruses, and cells. Cell 185, 2708–2724 (2022).

    Article  Google Scholar 

  23. Chan, L. Y., Kosuri, S. & Endy, D. Refactoring bacteriophage T7. Mol. Syst. Biol. 1, 2005.0018 (2005).

    Article  Google Scholar 

  24. Thi Nhu Thao, T. et al. Rapid reconstruction of SARS-CoV-2 using a synthetic genomics platform. Nature 582, 561–565 (2020).

    Article  Google Scholar 

  25. Dormitzer, P. R. et al. Synthetic generation of influenza vaccine viruses for rapid response to pandemics. Sci. Transl. Med. 5, 185ra68 (2013).

    Article  Google Scholar 

  26. Gibson, D. G. et al. Complete chemical synthesis, assembly, and cloning of a Mycoplasma genitalium genome. Science 319, 1215–1220 (2008).

    Article  Google Scholar 

  27. Pelletier, J. F. et al. Genetic requirements for cell division in a genomically minimal cell. Cell 184, 2430–2440.e16 (2021).

    Article  Google Scholar 

  28. Ostrov, N. et al. Design, synthesis, and testing toward a 57-codon genome. Science 353, 819–822 (2016).

    Article  Google Scholar 

  29. Wang, K. et al. Defining synonymous codon compression schemes by genome recoding. Nature 539, 59–64 (2016).

    Article  Google Scholar 

  30. Goffeau, A. et al. Life with 6000 genes. Science 274, 546–567 (1996).

    Article  Google Scholar 

  31. Mitchell, L. A. et al. Synthesis, debugging, and effects of synthetic chromosome consolidation: synVI and beyond. Science 355, eaaf4831 (2017).

    Article  Google Scholar 

  32. Shen, Y. et al. Deep functional analysis of synII, a 770-kilobase synthetic yeast chromosome. Science 355, eaaf4791 (2017).

    Article  Google Scholar 

  33. Wu, Y. et al. Bug mapping and fitness testing of chemically synthesized chromosome X. Science 355, eaaf4706 (2017).

    Article  Google Scholar 

  34. Xie, Z.-X. et al. ‘Perfect’ designer chromosome V and behavior of a ring derivative. Science 355, eaaf4704 (2017).

    Article  Google Scholar 

  35. Zhang, W. et al. Engineering the ribosomal DNA in a megabase synthetic chromosome. Science 355, eaaf3981 (2017).

    Article  Google Scholar 

  36. Zhao, Y. et al. Debugging and consolidating multiple synthetic chromosomes reveals combinatorial genetic interactions. Cell 186, 5220–5236.e16 (2023).

    Article  Google Scholar 

  37. Schindler, D. et al. Design, construction, and functional characterization of a tRNA neochromosome in yeast. Cell 186, 5237–5253.e22 (2023).

    Article  Google Scholar 

  38. Admire, A. et al. Cycles of chromosome instability are associated with a fragile site and are increased by defects in DNA replication and checkpoint controls in yeast. Genes Dev. 20, 159–173 (2006).

    Article  Google Scholar 

  39. Hamperl, S., Bocek, M. J., Saldivar, J. C., Swigut, T. & Cimprich, K. A. Transcription–replication conflict orientation modulates R-loop levels and activates distinct DNA damage responses. Cell 170, 774–786.e19 (2017).

    Article  Google Scholar 

  40. Mularoni, L. et al. Retrotransposon Ty1 integration targets specifically positioned asymmetric nucleosomal DNA segments in tRNA hotspots. Genome Res. 22, 693–703 (2012).

    Article  Google Scholar 

  41. Dai, J., Boeke, J. D., Luo, Z., Jiang, S. & Cai, Y. Sc3.0: revamping and minimizing the yeast genome. Genome Biol. 21, 205 (2020).

    Article  Google Scholar 

  42. Birchler, J. A., Graham, N. D., Swyers, N. C., Cody, J. P. & McCaw, M. E. Plant minichromosomes. Curr. Opin. Biotechnol. 37, 135–142 (2016).

    Article  Google Scholar 

  43. Shay, J. W. & Wright, W. E. Telomeres and telomerase: three decades of progress. Nat. Rev. Genet. 20, 299–309 (2019).

    Article  Google Scholar 

  44. Yu, W., Lamb, J. C., Han, F. & Birchler, J. A. Telomere-mediated chromosomal truncation in maize. Proc. Natl Acad. Sci. USA 103, 17331–17336 (2006).

    Article  Google Scholar 

  45. Carlson, W. R. & Phillips, R. L. The B chromosome of maize. Crit. Rev. Plant Sci. 3, 201–226 (1986).

    Article  Google Scholar 

  46. Gaeta, R. T. et al. In vivo modification of a maize engineered minichromosome. Chromosoma 122, 221–232 (2013).

    Article  Google Scholar 

  47. Teo, C. H. et al. Induction of telomere-mediated chromosomal truncation and stability of truncated chromosomes in Arabidopsis thaliana. Plant J. 68, 28–39 (2011).

    Article  Google Scholar 

  48. Murata, M. Minichromosomes and artificial chromosomes in Arabidopsis. Chr. Res. 22, 167–178 (2014).

    Article  Google Scholar 

  49. Xu, C., Cheng, Z. & Yu, W. Construction of rice mini-chromosomes by telomere-mediated chromosomal truncation. Plant J. Cell Mol. Biol. 70, 1070–1079 (2012).

    Article  Google Scholar 

  50. Kapusi, E. et al. Telomere-mediated truncation of barley chromosomes. Chromosoma 121, 181–190 (2012).

    Article  Google Scholar 

  51. Kouprina, N. et al. Human artificial chromosome with regulated centromere: a tool for genome and cancer studies. ACS Synth. Biol. 7, 1974–1989 (2018).

    Article  Google Scholar 

  52. Jiang, S. et al. Efficient de novo assembly and modification of large DNA fragments. Sci. China Life Sci. 65, 1445–1455 (2022).

    Article  Google Scholar 

  53. Cove, D. The moss Physcomitrella patens. Annu. Rev. Genet. 39, 339–358 (2005).

    Article  Google Scholar 

  54. Rensing, S. A., Goffinet, B., Meyberg, R., Wu, S.-Z. & Bezanilla, M. The moss Physcomitrium (Physcomitrella) patens: a model organism for non-seed plants. Plant Cell 32, 1361–1376 (2020).

    Article  Google Scholar 

  55. de Keijzer, J., Freire Rios, A. & Willemsen, V. Physcomitrium patens: a single model to study oriented cell divisions in 1D to 3D patterning. Int. J. Mol. Sci. 22, 2626 (2021).

    Article  Google Scholar 

  56. Bi, G. et al. Near telomere-to-telomere genome of the model plant Physcomitrium patens. Nat. Plants 10, 327–343 (2024).

    Article  Google Scholar 

  57. Schaefer, D. G. & Zrÿd, J.-P. Efficient gene targeting in the moss Physcomitrella patens. Plant J. 11, 1195–1206 (1997).

    Article  Google Scholar 

  58. Charlot, F., Goudounet, G., Nogué, F. & Perroud, P.-F. in Protoplast Technology: Methods and Protocols (eds Wang, K. & Zhang, F.) 3–19 (Humana Press, 2022).

  59. Widiez, T. et al. The chromatin landscape of the moss Physcomitrella patens and its dynamics during development and drought stress. Plant J. 79, 67–81 (2014).

    Article  Google Scholar 

  60. Lang, D. et al. The Physcomitrella patens chromosome-scale assembly reveals moss genome structure and evolution. Plant J. 93, 515–533 (2018).

    Article  Google Scholar 

  61. Khairul Ikram, N. K., Zakariya, A. M., Saiman, M. Z., Kashkooli, A. B. & Simonsen, H. T. Heterologous production of artemisinin in Physcomitrium patens by direct in vivo assembly of multiple DNA fragments. Bio-Protocol 13, e4719 (2023).

    Google Scholar 

  62. Reski, R., Parsons, J. & Decker, E. L. Moss-made pharmaceuticals: from bench to bedside. Plant Biotechnol. J. 13, 1191–1198 (2015).

    Article  Google Scholar 

  63. Decker, E. L., Parsons, J. & Reski, R. Glyco-engineering for biopharmaceutical production in moss bioreactors. Front. Plant Sci. 5, 346 (2014).

    Article  Google Scholar 

  64. Yu, W. et al. Designing a synthetic moss genome using GenoDesigner. Nat. Plants 10, 848–856 (2024). This study reports a systematic framework to govern synthesized chromosome in Physcomitrella patens, integrating sequence modularization and orthogonal design constraints.

    Article  Google Scholar 

  65. Gao, D. Introduction of plant transposon annotation for beginners. Biology 12, 1468 (2023).

    Article  Google Scholar 

  66. Elliott, T. A., Linquist, S. & Gregory, T. R. Conceptual and empirical challenges of ascribing functions to transposable elements. Am. Nat. 184, 14–24 (2014).

    Article  Google Scholar 

  67. Doolittle, W. F., Brunet, T. D. P., Linquist, S. & Gregory, T. R. Distinguishing between ‘function’ and ‘effect’ in genome biology. Genome Biol. Evol. 6, 1234–1237 (2014).

    Article  Google Scholar 

  68. Gemmell, N. J. Repetitive DNA: genomic dark matter matters. Nat. Rev. Genet. 22, 342 (2021).

    Article  Google Scholar 

  69. Buckley, P. T., Khaladkar, M., Kim, J. & Eberwine, J. Cytoplasmic intron retention, function, splicing, and the sentinel RNA hypothesis. Wiley Interdiscip. Rev. RNA 5, 223–230 (2014).

    Article  Google Scholar 

  70. Schmitz, U. et al. Intron retention enhances gene regulatory complexity in vertebrates. Genome Biol. 18, 216 (2017).

    Article  Google Scholar 

  71. Shaul, O. How introns enhance gene expression. Int. J. Biochem. Cell Biol. 91, 145–155 (2017).

    Article  Google Scholar 

  72. Erikson, O., Hertzberg, M. & Näsholm, T. A conditional marker gene allowing both positive and negative selection in plants. Nat. Biotechnol. 22, 455–458 (2004).

    Article  Google Scholar 

  73. Dymond, J. & Boeke, J. The Saccharomyces cerevisiae SCRaMbLE system and genome minimization. Bioeng. Bugs 3, 168–171 (2012).

    Google Scholar 

  74. Comai, L., Maheshwari, S. & Marimuthu, M. P. A. Plant centromeres. Curr. Opin. Plant Biol. 36, 158–167 (2017).

    Article  Google Scholar 

  75. Melters, D. P. et al. Comparative analysis of tandem repeats from hundreds of species reveals unique insights into centromere evolution. Genome Biol. 14, R10 (2013).

    Article  Google Scholar 

  76. McKinley, K. L. & Cheeseman, I. M. The molecular basis for centromere identity and function. Nat. Rev. Mol. Cell Biol. 17, 16–29 (2016).

    Article  Google Scholar 

  77. Carroll, C. W., Milks, K. J. & Straight, A. F. Dual recognition of CENP-A nucleosomes is required for centromere assembly. J. Cell Biol. 189, 1143–1155 (2010).

    Article  Google Scholar 

  78. Harrington, J. J., Bokkelen, G. V., Mays, R. W., Gustashaw, K. & Willard, H. F. Formation of de novo centromeres and construction of first-generation human artificial microchromosomes. Nat. Genet. 15, 345–355 (1997).

    Article  Google Scholar 

  79. Ebersole, T. et al. Rapid generation of long synthetic tandem repeats and its application for analysis in human artificial chromosome formation. Nucleic Acids Res. 33, e130 (2005).

    Article  Google Scholar 

  80. Noskov, V. N., Lee, N. C., Larionov, V. & Kouprina, N. Rapid generation of long tandem DNA repeat arrays by homologous recombination in yeast to study their function in mammalian genomes. Biol. Proc. Online 13, 8 (2011).

    Article  Google Scholar 

  81. Nakano, M. et al. Inactivation of a human kinetochore by specific targeting of chromatin modifiers. Dev. Cell 14, 507–522 (2008).

    Article  Google Scholar 

  82. Kouprina, N. et al. Organization of synthetic alphoid DNA array in human artificial chromosome (HAC) with a conditional centromere. ACS Synth. Biol. 1, 590–601 (2012).

    Article  Google Scholar 

  83. Logsdon, G. A. et al. Human artificial chromosomes that bypass centromeric DNA. Cell 178, 624–639.e19 (2019).

    Article  Google Scholar 

  84. Phan, B. H. et al. Transformation of rice with long DNA-segments consisting of random genomic DNA or centromere-specific DNA. Transgenic Res. 16, 341–351 (2007).

    Article  Google Scholar 

  85. Dawe, R. K. Charting the path to fully synthetic plant chromosomes. Exp. Cell Res. 390, 111951 (2020).

    Article  Google Scholar 

  86. Gent, J. I., Dong, Y., Jiang, J. & Dawe, R. K. Strong epigenetic similarity between maize centromeric and pericentromeric regions at the level of small RNAs, DNA methylation and H3 chromatin modifications. Nucleic Acids Res. 40, 1550–1560 (2012).

    Article  Google Scholar 

  87. Teo, C. H., Lermontova, I., Houben, A., Mette, M. F. & Schubert, I. De novo generation of plant centromeres at tandem repeats. Chromosoma 122, 233–241 (2013).

    Article  Google Scholar 

  88. Dawe, R. K. et al. Synthetic maize centromeres transmit chromosomes across generations. Nat. Plants 9, 433–441 (2023). This article describes the engineering of LexA–CENH3 chimaeras to mediate accurate recruitment of centromeric histones to LexO repeats, establishing de novo kinetochore complexes that triggered programmable chromosome breakage.

    Article  Google Scholar 

  89. Zhang, H. & Dawe, R. K. Total centromere size and genome size are strongly correlated in ten grass species. Chromosome Res. 20, 403–412 (2012).

    Article  Google Scholar 

  90. Plačková, K., Bureš, P. & Zedek, F. Centromere size scales with genome size across eukaryotes. Sci. Rep. 11, 19811 (2021).

    Article  Google Scholar 

  91. Kumawat, S. & Choi, J. Y. No end in sight: mysteries of the telomeric variation in plants. Am. J. Bot. 110, e16244 (2023).

    Article  Google Scholar 

  92. Zvereva, M. I., Shcherbakova, D. M. & Dontsova, O. A. Telomerase: structure, functions, and activity regulation. Biochemistry (Mosc) 75, 1563–1583 (2010).

    Google Scholar 

  93. Heacock, M., Spangler, E., Riha, K., Puizina, J. & Shippen, D. E. Molecular analysis of telomere fusions in Arabidopsis: multiple pathways for chromosome end‐joining. EMBO J. 23, 2304–2313 (2004).

    Article  Google Scholar 

  94. Kosuri, S. & Church, G. M. Large-scale de novo DNA synthesis: technologies and applications. Nat. Methods 11, 499–507 (2014).

    Article  Google Scholar 

  95. Beaucage, S. L. & Caruthers, M. H. Deoxynucleoside phosphoramidites — a new class of key intermediates for deoxypolynucleotide synthesis. Tetrahedron Lett. 22, 1859–1862 (1981).

    Article  Google Scholar 

  96. LeProust, E. M. et al. Synthesis of high-quality libraries of long (150mer) oligonucleotides by a novel depurination controlled process. Nucleic Acids Res. 38, 2522–2540 (2010).

    Article  Google Scholar 

  97. Matzas, M. et al. High-fidelity gene synthesis by retrieval of sequence-verified DNA identified using high-throughput pyrosequencing. Nat. Biotechnol. 28, 1291–1294 (2010).

    Article  Google Scholar 

  98. Palluk, S. et al. De novo DNA synthesis using polymerase–nucleotide conjugates. Nat. Biotechnol. 36, 645–650 (2018).

    Article  Google Scholar 

  99. Barthel, S., Palluk, S., Hillson, N. J., Keasling, J. D. & Arlow, D. H. Enhancing terminal deoxynucleotidyl transferase activity on substrates with 3′ terminal structures for enzymatic de novo DNA synthesis. Genes 11, 102 (2020).

    Article  Google Scholar 

  100. Benoit, R. M. et al. Seamless insert-plasmid assembly at high efficiency and low cost. PLoS ONE 11, e0153158 (2016).

    Article  Google Scholar 

  101. Gibson, D. G. et al. Enzymatic assembly of DNA molecules up to several hundred kilobases. Nat. Methods 6, 343–345 (2009).

    Article  Google Scholar 

  102. Engler, C., Gruetzner, R., Kandzia, R. & Marillonnet, S. Golden Gate shuffling: a one-pot DNA shuffling method based on type IIs restriction enzymes. PLOS ONE 4, e5553 (2009).

    Article  Google Scholar 

  103. Gibson, D. G. Gene and genome construction in yeast. Curr. Protoc. Mol. Biol. Ch. 3, Unit3.22 (2011).

    Google Scholar 

  104. Muller, H. et al. Assembling large DNA segments in yeast. Methods Mol. Biol. 852, 133–150 (2012).

    Article  Google Scholar 

  105. Gelvin, S. B. Integration of Agrobacterium T-DNA into the plant genome. Annu. Rev. Genet. 51, 195–217 (2017).

    Article  Google Scholar 

  106. Hwang, H.-H., Yu, M. & Lai, E.-M. Agrobacterium-mediated plant transformation: biology and applications. Arab. Book 15, e0186 (2017).

    Article  Google Scholar 

  107. Altpeter, F. et al. Particle bombardment and the genetic enhancement of crops: myths and realities. Mol. Breed. 15, 305–327 (2005).

    Article  Google Scholar 

  108. Lowe, B. A. et al. Enhanced single copy integration events in corn via particle bombardment using low quantities of DNA. Transgenic Res. 18, 831–840 (2009).

    Article  Google Scholar 

  109. Hamilton, C. M., Frary, A., Lewis, C. & Tanksley, S. D. Stable transfer of intact high molecular weight DNA into plant chromosomes. Proc. Natl Acad. Sci. USA 93, 9975–9979 (1996).

    Article  Google Scholar 

  110. Liu, J. et al. Genome-scale sequence disruption following biolistic transformation in rice and maize. Plant Cell 31, 368–383 (2019).

    Article  Google Scholar 

  111. Ikeuchi, M., Sugimoto, K. & Iwase, A. Plant callus: mechanisms of induction and repression. Plant Cell 25, 3159–3173 (2013).

    Article  Google Scholar 

  112. Duarte, P., Ribeiro, D., Carqueijeiro, I., Bettencourt, S. & Sottomayor, M. in Biotechnology of Plant Secondary Metabolism: Methods and Protocols (ed. Fett-Neto, A. G.) 137–148 (Humana Press, 2016).

  113. Li, L. & Blankenstein, T. Generation of transgenic mice with megabase-sized human yeast artificial chromosomes by yeast spheroplast–embryonic stem cell fusion. Nat. Protoc. 8, 1567–1582 (2013).

    Article  Google Scholar 

  114. Brown, D. M. et al. Efficient size-independent chromosome delivery from yeast to cultured cell lines. Nucleic Acids Res. 45, e50–e50 (2017).

    Google Scholar 

  115. Wu, F.-H., Yuan, Y.-H., Hsu, C.-T., Cheng, Q.-W. & Lin, C.-S. in Protoplast Technology: Methods and Protocols (eds Wang, K. & Zhang, F.) 49–64 (Humana Press, 2022).

  116. Jones, A. M. P., Shukla, M. R., Biswas, G. C. G. & Saxena, P. K. Protoplast-to-plant regeneration of American elm (Ulmus americana). Protoplasma 252, 925–931 (2015).

    Article  Google Scholar 

  117. Jeong, Y. Y., Lee, H.-Y., Kim, S. W., Noh, Y.-S. & Seo, P. J. Optimization of protoplast regeneration in the model plant Arabidopsis thaliana. Plant Methods 17, 21 (2021).

    Article  Google Scholar 

  118. Lowe, K. et al. Morphogenic regulators Baby boom and Wuschel improve monocot transformation. Plant Cell 28, 1998–2015 (2016).

    Article  Google Scholar 

  119. Debernardi, J. M. et al. A GRF–GIF chimeric protein improves the regeneration efficiency of transgenic plants. Nat. Biotechnol. 38, 1274–1279 (2020).

    Article  Google Scholar 

  120. Yu, Y. et al. Enhancing wheat regeneration and genetic transformation through overexpression of TaLAX1. Focus Issue Wheat Biol. 5, 100738 (2024).

    Google Scholar 

  121. Liu, X. et al. Uncovering the transcriptional regulatory network involved in boosting wheat regeneration and transformation. Nat. Plants 9, 908–925 (2023).

    Article  Google Scholar 

  122. Xu, M., Du, Q., Tian, C., Wang, Y. & Jiao, Y. Stochastic gene expression drives mesophyll protoplast regeneration. Sci. Adv. 7, eabg8466 (2021).

    Article  Google Scholar 

  123. Yang, W. et al. Peptide REF1 is a local wound signal promoting plant regeneration. Cell 187, 3024–3038.e14 (2024).

    Article  Google Scholar 

  124. Wang, N. et al. Leaf transformation for efficient random integration and targeted genome modification in maize and sorghum. Nat. Plants 9, 255–270 (2023).

    Google Scholar 

  125. Sun, Y. et al. Engineering herbicide-resistant rice plants through CRISPR/Cas9-mediated homologous recombination of acetolactate synthase. Mol. Plant 9, 628–631 (2016).

    Article  Google Scholar 

  126. Schmidt, C., Pacher, M. & Puchta, H. in Transgenic Plants: Methods and Protocols (eds Kumar, S. et al.) 237–266 (Humana Press, 2019).

  127. Fauser, F. et al. In planta gene targeting. Proc. Natl Acad. Sci. USA 109, 7535–7540 (2012).

    Article  Google Scholar 

  128. Mali, P. et al. RNA-guided human genome engineering via Cas9. Science 339, 823–826 (2013).

    Article  Google Scholar 

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

    Article  Google Scholar 

  130. Zhao, Y. et al. An alternative strategy for targeted gene replacement in plants using a dual-sgRNA/Cas9 design. Sci. Rep. 6, 23890 (2016).

    Article  Google Scholar 

  131. Rees, H. A., Yeh, W.-H. & Liu, D. R. Development of hRad51–Cas9 nickase fusions that mediate HDR without double-stranded breaks. Nat. Commun. 10, 2212 (2019).

    Article  Google Scholar 

  132. Chauhan, V. P., Sharp, P. A. & Langer, R. Altered DNA repair pathway engagement by engineered CRISPR–Cas9 nucleases. Proc. Natl Acad. Sci. USA 120, e2300605120 (2023).

    Article  Google Scholar 

  133. Schreiber, T. et al. Efficient scar-free knock-ins of several kilobases in plants by engineered CRISPR–Cas endonucleases. Mol. Plant 17, 824–837 (2024).

    Article  Google Scholar 

  134. Ali, Z. et al. Fusion of the Cas9 endonuclease and the VirD2 relaxase facilitates homology-directed repair for precise genome engineering in rice. Commun. Biol. 3, 44 (2020).

    Article  Google Scholar 

  135. Morton, J., Davis, M. W., Jorgensen, E. M. & Carroll, D. Induction and repair of zinc-finger nuclease-targeted double-strand breaks in Caenorhabditis elegans somatic cells. Proc. Natl Acad. Sci. USA 103, 16370–16375 (2006).

    Article  Google Scholar 

  136. Maruyama, T. et al. Increasing the efficiency of precise genome editing with CRISPR–Cas9 by inhibition of nonhomologous end joining. Nat. Biotechnol. 33, 538–542 (2015).

    Article  Google Scholar 

  137. Qi, Y. et al. Increasing frequencies of site-specific mutagenesis and gene targeting in Arabidopsis by manipulating DNA repair pathways. Genome Res. 23, 547–554 (2013).

    Article  Google Scholar 

  138. Song, J. et al. RS-1 enhances CRISPR/Cas9- and TALEN-mediated knock-in efficiency. Nat. Commun. 7, 10548 (2016).

    Article  Google Scholar 

  139. Karasu, M. E. et al. Removal of TREX1 activity enhances CRISPR–Cas9-mediated homologous recombination. Nat. Biotechnol. https://doi.org/10.1038/s41587-024-02356-3 (2024).

    Article  Google Scholar 

  140. Lu, Y. et al. Targeted, efficient sequence insertion and replacement in rice. Nat. Biotechnol. 38, 1402–1407 (2020).

    Article  Google Scholar 

  141. Liu, P. et al. Transposase-assisted target-site integration for efficient plant genome engineering. Nature 631, 593–600 (2024).

    Article  Google Scholar 

  142. Kühn, R. & Torres, R. M. in Transgenesis Techniques: Principles and Protocols (ed. Clarke, A. R.) 175–204 (Springer, 2002).

  143. Sun, C. et al. Precise integration of large DNA sequences in plant genomes using PrimeRoot editors. Nat. Biotechnol. 42, 316–327 (2024).

    Article  Google Scholar 

  144. Rönspies, M., Dorn, A., Schindele, P. & Puchta, H. CRISPR–Cas-mediated chromosome engineering for crop improvement and synthetic biology. Nat. Plants 7, 566–573 (2021).

    Article  Google Scholar 

  145. Rönspies, M., Schindele, P., Wetzel, R. & Puchta, H. CRISPR–Cas9-mediated chromosome engineering in Arabidopsis thaliana. Nat. Protoc. 17, 1332–1358 (2022).

    Article  Google Scholar 

  146. Schmidt, C. et al. Changing local recombination patterns in Arabidopsis by CRISPR/Cas mediated chromosome engineering. Nat. Commun. 11, 4418 (2020).

    Article  Google Scholar 

  147. Rönspies, M. et al. Massive crossover suppression by CRISPR–Cas-mediated plant chromosome engineering. Nat. Plants 8, 1153–1159 (2022).

    Article  Google Scholar 

  148. Beying, N., Schmidt, C., Pacher, M., Houben, A. & Puchta, H. CRISPR–Cas9-mediated induction of heritable chromosomal translocations in Arabidopsis. Nat. Plants 6, 638–645 (2020).

    Article  Google Scholar 

  149. Godwin, J. & Farrona, S. Plant epigenetic stress memory induced by drought: a physiological and molecular perspective. Methods Mol. Biol. 2093, 243–259 (2020).

    Article  Google Scholar 

  150. Hemenway, E. A. & Gehring, M. Epigenetic regulation during plant development and the capacity for epigenetic memory. Annu. Rev. Plant Biol. 74, 87–109 (2023).

    Article  Google Scholar 

  151. Gjaltema, R. A. F. & Rots, M. G. Advances of epigenetic editing. Curr. Opin. Chem. Biol. 57, 75–81 (2020).

    Article  Google Scholar 

  152. Srivastava, V. & Thomson, J. Gene stacking by recombinases. Plant Biotechnol. J. 14, 471–482 (2016).

    Article  Google Scholar 

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Acknowledgements

Work in the authors’ laboratory is funded by the National Key R&D Program of China (2023YFE0101100 and 2019YFA0903900), the Natural Science Foundation of China (32230010 and 32270345), the Qidong-SLS Innovation Fund, and the Guangdong Provincial Key Laboratory of Synthetic Genomics (2023B1212060054).

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The manuscript was written by T.L. All the authors contributed to discussion of the content and review and editing of the manuscript.

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Correspondence to Ying Wang  (汪颖) or Yuling Jiao  (焦雨铃).

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Lan, T., Chen, LG., Wang, Y. et al. Genome synthesis in plants. Nat Rev Bioeng (2025). https://doi.org/10.1038/s44222-025-00326-1

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