| Abstract [eng] |
Recombinant proteins are already successfully produced in plants in contained facilities using Agrobacterium-mediated transient transfection of plants. However, transfection methods suitable for open field applications are still desirable as a cheaper alternative. Biosafety concerns related to the use of recombinant agrobacteria in an industrial transfection process include possible transformation/transfection of unintended hosts or spread of the genetically modified agrobacteria in the environment. In this work, we explored novel biocontrol approaches resulting in higher biosafety of the transient expression in plants. Our proposed solutions involve an inducible expression of essential Agrobacterium tumefaciens virulence factor virE2 and expression of toxin pemK and antitoxin pemI from bacterial toxin-antitoxin (TA) module pemIK in Agrobacterium that provides for strictly regulated T-DNA transfer from agrobacteria to model plants Nicotiana benthamiana. The study provided detailed information about the relative activity and regulation of several IPTG and cumic acid inducible promoters in A. tumefaciens. Additionally, new efficient cumic acid inducible promoters were constructed. Also, several A. tumefaciens toxins were identified from putative TA systems, and their activity was evaluated. Toxins Dead, PIN, VapC, IetS and PemK showed high efficiency against A. tumefaciens. |
