The Mechanism of T-DNA Integration: Some Major Unresolved Questions.

The mechanism of T-DNA integration into plant genomes during Agrobacterium-mediated genetic transformation is still not understood. As genetic transformation of plants via Agrobacterium has become a routine practice among plant biologists, understanding T-DNA integration remains important for several reasons. First, T-DNA is the final step in one of the unique cases of inter-kingdom horizontal gene transfer in nature. Second, understanding T-DNA integration is important for biotechnological applications. For example, better knowledge of this process may help develop methods to transform species that are currently not susceptible to Agrobacterium-mediated transformation. In addition, regulatory agencies usually require "clean" and "precise" transgenic insertion events, whereas transgenic insertions are commonly complex unpredictable structures. Furthermore, whereas T-DNA integration under natural conditions occurs randomly, technology to direct T-DNA to specific sites in the genome is highly desired. A better understanding of T-DNA integration may help develop methods to achieve more desirable results. Finally, gene targeting methods that require a foreign DNA template for precise DNA modifications in plants often utilize Agrobacterium to deliver the DNA template. Better understanding of the fate of T-DNA in the plant nucleus may help utilize T-DNA for more efficient gene targeting. For introducing gene targeting reagents, efficient delivery of T-DNA without ectopic integration would be useful. The following review summarizes current knowledge related to T-DNA integration. Five major open questions related to T-DNA integration are being presented. Finally, different models for T-DNA integration are being discussed, and a revised model is proposed.