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Enhanced production of single copy backbone-free transgenic plants in multiple crop species using binary vectors with a pRi replication origin in Agrobacterium tumefaciens

Abstract

Single transgene copy, vector backbone-free transgenic crop plants are highly desired for functional genomics and many biotechnological applications. We demonstrate that binary vectors that use a replication origin derived from the Ri plasmid of Agrobacterium rhizogenes (oriRi) increase the frequency of single copy, backbone-free transgenic plants in Agrobacterium tumefaciens mediated transformation of soybean, canola, and corn, compared to RK2-derived binary vectors (RK2 oriV). In large scale soybean transformation experiments, the frequency of single copy, backbone-free transgenic plants was nearly doubled in two versions of the oriRi vectors compared to the RK2 oriV control vector. In canola transformation experiments, the oriRi vector produced more single copy, backbone-free transgenic plants than did the RK2 oriV vector. In corn transformation experiments, the frequency of single copy backbone-free transgenic plants was also significantly increased when using the oriRi vector, although the transformation frequency dropped. These results, derived from transformation experiments using three crops, indicate the advantage of oriRi vectors over RK2 oriV binary vectors for the production of single copy, backbone-free transgenic plants using Agrobacterium-mediated transformation.

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Acknowledgments

We thank many Monsanto colleagues for assistance during the course of the experiments and data analysis. Particularly, we thank Middleton Soybean Transformation Team for the large scale soybean transformation experiments; J. Rout, M. McKenzie, M. Mann, B. Davis and J. Kumpf at Monsanto Mystic Campus for corn plant transformation; Monsanto Trait Development teams at Middleton, Davis and Mystic for plant care in greenhouses; Monsanto Gene Assessment and Molecular Analysis teams at Middleton, St. Louis and Mystic for transgene copy number and other molecular analyses; Drs. D. Somers and B. Tinland for critical reading manuscript, and J. Harrison and C. Marquez for statistical analysis.

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Correspondence to Xudong Ye.

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Ye, X., Williams, E.J., Shen, J. et al. Enhanced production of single copy backbone-free transgenic plants in multiple crop species using binary vectors with a pRi replication origin in Agrobacterium tumefaciens . Transgenic Res 20, 773–786 (2011). https://doi.org/10.1007/s11248-010-9458-6

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Keywords

  • Agrobacterium-mediated transformation
  • Transgenic plants
  • Binary vector
  • Copy number
  • Single copy backbone-free transgenic plant
  • Soybean transformation
  • Canola transformation
  • Corn transformation