GENE ASSEMBLY IN AGROBACTERIUM VIA NUCLEIC ACID TRANSFER USING RECOMBINASE TECHNOLOGY (GAANTRY)
DOI:
https://doi.org/10.1808/8txehn94Keywords:
Plant biotechnology, Transformation, Agrobacterium rhizogenes, Transfer DNA (T-DNA), Transgene assembly, Site-specific recombination, Gene stacking, Virulence plasmid, Genetic engineeringAbstract
Plant biotechnology provides a means for the rapid genetic improvement of crops including the enhance ment of complex traits like yield and nutritional quality through the introduction and coordinated expression of multiple genes. GAANTRY (geneassembly in Agrobacterium bynucleic acidtransfer using recombinase technology) is a flexible and effective system for stably stacking multiple genes within an Agrobacterium virulence plasmid transfer DNA (T-DNA) region. The system provides a simple andefficient method for assembling and stably maintaining large stacked constructs within the GAANTRY ArPORT1 Agrobacterium rhizogenes strain. The assembly process utilizes unidirectional site-specific recombinases in vivo and an alternating bacterial selection scheme to sequentially assemble multiple genes into a single transformation construct. A detailed description of the procedures used for bacterial transformation, selection, counter selection, and genomic PCR validation with the GAANTRY system are presented. The methods described facilitate the efficient assembly and validation of large GAANTRY T-DNA constructs. This powerful, yet simple to use, technology will be a convenient tool for transgene stacking and plant genetic engineering of rice and other crop plants.
References
Gelvin SB (2003) Agrobacterium-mediated plant transformation: the biology behind the “gene-jockeying” tool. Microbiol Mol Biol Rev 67(1):16–37
Vain P (2007) Thirty years of plant transforma tion technology development. Plant Biotech nol J 5(2):221–229. https://doi.org/10. 1111/j.1467-7652.2006.00225.x
Dafny-Yelin M, Tzfira T (2007) Delivery of multiple transgenes to plant cells. Plant Physiol 145(4):1118–1128. https://doi.org/10. 1104/pp.107.106104
MaL,DongJ,JinY,ChenM,ShenX,WangT (2011) RMDAP: a versatile, ready-to-use tool box for multigene genetic transformation. PLoS One 6(5):e19883. https://doi.org/10. 1371/journal.pone.0019883
Weber E, Engler C, Gruetzner R, Werner S, Marillonnet S (2011) A modular cloning system for standardized assembly of multigene constructs. PLoS One 6(2):e16765. https:// doi.org/10.1371/journal.pone.0016765
Untergasser A, Bijl GJM, Liu W, Bisseling T, Schaart JG, Geurts R (2012) One-step Agro bacterium mediated transformation of eight genes essential for Rhizobium symbiotic signal ing using the novel binary vector system pHUGE. PLoS One 7(10):e47885. https:// doi.org/10.1371/journal.pone.0047885
Zeevi V, Liang Z, Arieli U, Tzfira T (2012) Zinc f inger nuclease and homing endonuclease-mediated assembly of multigene plant transformation vectors. Plant Physiol 158 (1):132–144. https://doi.org/10.1104/pp. 111.184374
Buntru M, G€ artner S, Staib L, Kreuzaler F, Schlaich N (2013) Delivery of multiple trans genes to plant cells by an improved version of GAANTRY Gene Stacking 17 MultiRound Gateway technology. Transgenic Res 22(1):153–167. https://doi.org/10. 1007/s11248-012-9640-0
Binder A, Lambert J, Morbitzer R, Popp C, Ott T,LahayeT,Parniske M(2014)Amodular 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
Shih PM, Vuu K, Mansoori N, Ayad L, Louie KB, Bowen BP, Northen TR, Loque´ D (2016) A robust gene-stacking method utilizing yeast assembly for plant synthetic biology. Nat Com mun 7:13215. https://doi.org/10.1038/ ncomms13215 C
Cermak T, Curtin SJ, Gil-Humanes J, egan R, Kono TJY, Konecˇna´ E, Belanto JJ, Starker CG, Mathre JW, Greenstein RL, Voytas DF (2017) A multi-purpose toolkit to enable advanced genome engineering in plants. Plant Cell. https://doi.org/10.1105/tpc.16.00922
Zhang H-Y, Wang X-H, Dong L, Wang Z-P, Liu B, Lv J, Xing H-L, Han C-Y, Wang X-C, Chen Q-J (2017) MISSA 2.0: an updated syn thetic biology toolbox for assembly of orthog onal CRISPR/Cas systems. Sci Rep 7:41993. https://doi.org/10.1038/srep41993
ZhuQ,YuS,ZengD,LiuH,WangH,YangZ, Xie X, Shen R, Tan J, Li H, Zhao X, Zhang Q, Chen Y, Guo J, Chen L, Liu Y-G (2017) Development of “purple endosperm rice” by engineering anthocyanin biosynthesis in the endosperm with a high-efficiency transgene stacking system. Mol Plant 10(7):918–929. https://doi.org/10.1016/j.molp.2017.05. 008
Collier R, Thomson JG, Thilmony R (2018) A versatile and robust Agrobacterium-based gene stacking system generates high-quality trans genic Arabidopsis plants. Plant J 95 (4):573–583. https://doi.org/10.1111/tpj. 13992
Green MR (2012) In: Green MR, Sambrook J (eds) Molecular cloning : a laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NYVolume accessed from https://nla.gov.au/nla.cat-vn6039452
Tu Q, Yin J, Fu J, Herrmann J, Li Y, Yin Y, Stewart AF, Muller R, Zhang Y (2016) Room temperature electrocompetent bacterial cells improve DNA transformation and recombi neering efficiency. Sci Rep 6:24648. https:// doi.org/10.1038/srep24648
Sallaud C, Meynard D, van Boxtel J, Gay C, Bes M, Brizard JP, Larmande P, Ortega D, Raynal M, Portefaix M, Ouwerkerk PB, Rueb S, Delseny M, Guiderdoni E (2003) Highly efficient production and characteriza tion of T-DNA plants for rice (Oryza sativa L.) functional genomics. Theor Appl Genet 106(8):1396–1408. https://doi.org/10. 1007/s00122-002-1184
