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The Use of an Automated Platform to Assemble Multigenic Constructs for Plant Transformation

  • David G. J. Mann
  • Scott A. Bevan
  • Anthony J. Harvey
  • Rachelle A. Leffert-Sorenson
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1864)

Abstract

Compared to traditional means, modern DNA assembly methods allow cloning of large, multigenic vectors for plant transformation in rapid fashion. These methods are often robust and efficient and can assemble multiple DNA fragments into a single vector in one reaction. Here we describe the use of an automated DNA assembly platform for the generation of complex, multigenic T-DNA binary vectors using a hierarchical Golden Gate cloning strategy. These DNA constructs contained diverse DNA elements for the expression of multiple genes for trait stacking in the crop of interest. This platform streamlines the DNA assembly and validation process through high-efficiency cloning methods, integrated automation equipment, and increased throughput. The implementation of this platform removes bottlenecks for routine molecular biology and opens new possibilities for downstream experimental idea testing.

Key words

DNA assembly Golden Gate cloning Plasmid T-DNA binary vectors Multigenic construct Plant biotechnology Synthetic biology Automation Liquid handler 

Notes

Acknowledgments

The authors would like to thank Bill Moskal for his intellectual input on the integration of an automated platform, Siobhan Davis for her collaboration on sample management and archiving, Ryan Blue and Patrick Westfall for their intellectual input on DNA assembly methods, and Ann Owens Merlo and Tom Meade for their leadership and support.

References

  1. 1.
    Gibson DG, Young L, Chuang RY, Venter JC, Hutchison CA 3rd, Smith HO (2009) Enzymatic assembly of DNA molecules up to several hundred kilobases. Nat Methods 6(5):343–345.  https://doi.org/10.1038/nmeth.1318 CrossRefPubMedGoogle Scholar
  2. 2.
    Juhas M, Ajioka J (2017) High molecular weight DNA assembly in vivo for synthetic biology applications. Crit Rev Biotechnol 37(3):277–286.  https://doi.org/10.3109/07388551.2016.1141394 CrossRefPubMedGoogle Scholar
  3. 3.
    Ellis T, Adie T, Baldwin GS (2011) DNA assembly for synthetic biology: from parts to pathways and beyond. Integr Biol (Camb) 3(2):109–118.  https://doi.org/10.1039/c0ib00070a CrossRefGoogle Scholar
  4. 4.
    Chao R, Liang J, Tasan I, Si T, Ju L, Zhao H (2017) Fully automated one-step synthesis of single-transcript TALEN pairs using a biological foundry. ACS Synth Biol 6(4):678–685.  https://doi.org/10.1021/acssynbio.6b00293 CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Kosuri S, Eroshenko N, LeProust EM, Super M, Way J, Li JB, Church GM (2010) Scalable gene synthesis by selective amplification of DNA pools from high-fidelity microchips. Nat Biotechnol 28(12):1295–1299.  https://doi.org/10.1038/nbt.1716 http://www.nature.com/nbt/journal/v28/n12/abs/nbt.1716.html#supplementary-information CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Engler C, Gruetzner R, Kandzia R, Marillonnet S (2009) Golden gate shuffling: a one-pot DNA shuffling method based on type IIs restriction enzymes. PLoS One 4(5):e5553.  https://doi.org/10.1371/journal.pone.0005553 CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Engler C, Kandzia R, Marillonnet S (2008) A one pot, one step, precision cloning method with high throughput capability. PLoS One 3(11):e3647.  https://doi.org/10.1371/journal.pone.0003647 CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Binder A, Lambert J, Morbitzer R, Popp C, Ott T, Lahaye T, Parniske M (2014) A modular plasmid assembly kit for multigene expression, gene silencing and silencing rescue in plants. PLoS One 9(2):e88218.  https://doi.org/10.1371/journal.pone.0088218 CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Boyle PM, Burrill DR, Inniss MC, Agapakis CM, Deardon A, Dewerd JG, Gedeon MA, Quinn JY, Paull ML, Raman AM, Theilmann MR, Wang L, Winn JC, Medvedik O, Schellenberg K, Haynes KA, Viel A, Brenner TJ, Church GM, Shah JV, Silver PA (2012) A BioBrick compatible strategy for genetic modification of plants. J Biol Eng 6(1):8.  https://doi.org/10.1186/1754-1611-6-8 CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Casini A, Storch M, Baldwin GS, Ellis T (2015) Bricks and blueprints: methods and standards for DNA assembly. Nat Rev Mol Cell Biol 16(9):568–576.  https://doi.org/10.1038/nrm4014 CrossRefPubMedGoogle Scholar
  11. 11.
    Engler C, Youles M, Gruetzner R, Ehnert T-M, Werner S, Jones JDG, Patron NJ, Marillonnet S (2014) A Golden Gate modular cloning toolbox for plants. ACS Synth Biol 3(11):839–843.  https://doi.org/10.1021/sb4001504 CrossRefPubMedGoogle Scholar
  12. 12.
    Guo Y, Dong J, Zhou T, Auxillos J, Li T, Zhang W, Wang L, Shen Y, Luo Y, Zheng Y, Lin J, Chen GQ, Wu Q, Cai Y, Dai J (2015) YeastFab: the design and construction of standard biological parts for metabolic engineering in Saccharomyces cerevisiae. Nucleic Acids Res 43(13):e88.  https://doi.org/10.1093/nar/gkv464 CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Patron NJ, Orzaez D, Marillonnet S, Warzecha H, Matthewman C, Youles M, Raitskin O, Leveau A, Farré G, Rogers C, Smith A, Hibberd J, Webb AAR, Locke J, Schornack S, Ajioka J, Baulcombe DC, Zipfel C, Kamoun S, Jones JDG, Kuhn H, Robatzek S, Van Esse HP, Sanders D, Oldroyd G, Martin C, Field R, O'Connor S, Fox S, Wulff B, Miller B, Breakspear A, Radhakrishnan G, Delaux P-M, Loqué D, Granell A, Tissier A, Shih P, Brutnell TP, Quick WP, Rischer H, Fraser PD, Aharoni A, Raines C, South PF, Ané J-M, Hamberger BR, Langdale J, Stougaard J, Bouwmeester H, Udvardi M, Murray JAH, Ntoukakis V, Schäfer P, Denby K, Edwards KJ, Osbourn A, Haseloff J (2015) Standards for plant synthetic biology: a common syntax for exchange of DNA parts. New Phytol 208(1):13–19.  https://doi.org/10.1111/nph.13532 CrossRefPubMedGoogle Scholar
  14. 14.
    Sarrion-Perdigones A, Vazquez-Vilar M, Palaci J, Castelijns B, Forment J, Ziarsolo P, Blanca J, Granell A, Orzaez D (2013) GoldenBraid 2.0: a comprehensive DNA assembly framework for plant synthetic biology. Plant Physiol 162(3):1618–1631.  https://doi.org/10.1104/pp.113.217661 CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Sarrion-Perdigones A, Palaci J, Granell A, Orzaez D (2014) Design and construction of multigenic constructs for plant biotechnology using the GoldenBraid cloning strategy. Methods Mol Biol 1116:133–151.  https://doi.org/10.1007/978-1-62703-764-8_10 CrossRefPubMedGoogle Scholar
  16. 16.
    Sarrion-Perdigones A, Falconi EE, Zandalinas SI, Juarez P, Fernandez-del-Carmen A, Granell A, Orzaez D (2011) GoldenBraid: an iterative cloning system for standardized assembly of reusable genetic modules. PLoS One 6:e21622CrossRefGoogle Scholar
  17. 17.
    Weber E, Engler C, Gruetzner R, Werner S, Marillonnet S (2011) A modular cloning system for standardized assembly of multigene constructs. PLoS One 6:e16765CrossRefGoogle Scholar
  18. 18.
    Chen J, Densmore D, Ham TS, Keasling JD, Hillson NJ (2012) DeviceEditor visual biological CAD canvas. J Biol Eng 6:1.  https://doi.org/10.1186/1754-1611-6-1 CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Galdzicki M, Clancy KP, Oberortner E, Pocock M, Quinn JY, Rodriguez CA (2014) The Synthetic Biology Open Language (SBOL) provides a community standard for communicating designs in synthetic biology. Nat Biotechnol 32:545.  https://doi.org/10.1038/nbt.2891 CrossRefPubMedGoogle Scholar
  20. 20.
    Gansner ER, North SC (2000) An open graph visualization system and its applications to software engineering. Softw Pract Exper 30(11):1203–1233CrossRefGoogle Scholar
  21. 21.
    Hillson NJ, Rosengarten RD, Keasling JD (2012) j5 DNA assembly design automation software. ACS Synth Biol 1(1):14–21.  https://doi.org/10.1021/sb2000116 CrossRefPubMedGoogle Scholar
  22. 22.
    Johnson James R, D'Amore R, Thain Simon C, Craig T, McCue Hannah V, Hertz-Fowler C, Hall N, Hall Anthony JW (2016) GeneMill: a 21st century platform for innovation. Biochem Soc Trans 44(3):681–683.  https://doi.org/10.1042/bst20160012 CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Linshiz G, Jensen E, Stawski N, Bi C, Elsbree N, Jiao H, Kim J, Mathies R, Keasling JD, Hillson NJ (2016) End-to-end automated microfluidic platform for synthetic biology: from design to functional analysis. J Biol Eng 10(1):3.  https://doi.org/10.1186/s13036-016-0024-5 CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Linshiz G, Stawski N, Goyal G, Bi C, Poust S, Sharma M (2014) PR-PR: cross-platform laboratory automation system. ACS Synth Biol 3:515.  https://doi.org/10.1021/sb4001728 CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • David G. J. Mann
    • 1
  • Scott A. Bevan
    • 1
  • Anthony J. Harvey
    • 1
  • Rachelle A. Leffert-Sorenson
    • 1
  1. 1.Corteva Agriscience™Agriculture Division of DowDuPont™IndianapolisUSA

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