Abstract
Agrobacterium tumefaciens has been widely used as a tool for transgenesis in plants. The availability of its genome sequence should facilitate the directed engineering of improved properties; however, the current genome engineering options are laborious. Here, we investigated whether the lambda R ed operon can be applied for recombineering of the A. tumefaciens genome. First, we built an expression plasmid for A. tumefaciens employing a tetracycline-inducible promoter to regulate the Red operon. This multicopy plasmid was then itself modified in A. tumefaciens to verify that the Red operon was functional. Then, we modified the endogenous A. tumefaciens tumor-inducing plasmid and the linear chromosome. These results extend recombineering technology to a new host and indicate a fast and convenient way to engineer the A. tumefaciens genome for functional genomics and strain improvements.
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Acknowledgments
This investigation was supported by the National Natural Science Foundation of China (31200004), the Hunan Provincial Natural Science Foundation of China (14JJ6009), the Specialized Research Fund for the Doctoral Program of Higher Education from the Ministry of Education of China (2011430620005), the Scientific Research Fund of Hunan Provincial Education Department (10K041).
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Hu, S., Fu, J., Huang, F. et al. Genome engineering of Agrobacterium tumefaciens using the lambda Red recombination system. Appl Microbiol Biotechnol 98, 2165–2172 (2014). https://doi.org/10.1007/s00253-013-5412-x
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DOI: https://doi.org/10.1007/s00253-013-5412-x