Identification of plant genes involved in Agrobacterium-mediated plant transformation

Agrobacterium tumefaciens is a Gram negative soil bacterium that can cause crown gall disease in plants. Genetic transformation of plant cells by A. tumefaciens represents a unique case of trans-kingdom sex requiring the involvement of both bacterial virulence proteins and plant encoded proteins. DNA transfer between A. tumefaciens and plants is the only known inter-kingdom natural DNA transfer. Agrobacterium-mediated plant transformation is the most common and preferred method to produce transgenic plants. In spite of significant progress in Agrobacterium-mediated plant transformation techniques, it is still not easy to genetically transform several agronomically important crops like corn and soybean using A. tumefaciens. A better understanding of this unique inter-kingdom DNA transfer between Agrobacterium and plants will help us improve the crop transformation efficiency and to prevent crown gall disease on economically important fruit crops like grapes.

We have developed in planta and leaf disk assays in Nicotiana benthamiana for identifying plant genes involved in Agrobacterium-mediated plant transformation using virus-induced gene silencing (VIGS) as a genomics tool. A reverse genetics approach based on VIGS was used to revalidate the role of several genes that are either known or speculated to be involved in Agrobacterium-mediated plant transformation. By employing the combination of VIGS-mediated forward genetics approach and an in planta tumorigenesis assay we identified several ACG (altered crown gall) genes that when silenced produced altered crown gall phenotypes upon infection with a tumorigenic Agrobacterium strain. The VIGS-based approach to identify and characterize plant genes involved in genetic transformation of plant cells by Agrobacterium is simple, rapid and robust, and complements other currently used approaches.

This project is funded by the Samuel Roberts Noble Foundation and NSF award IOB-0445799.