A repression system for genes encoding proteins toxic to bacteria in Agrobacterium tumefaciens-mediated plant transformation

Abstract

Transfer of a chimeric gene encoding secretory T4 lysozyme under the control of the mannopine synthase promoter (pMAS) into tobacco by Agrobacterium-mediated gene transfer has proven to be difficult. This can be due to unwanted expression of the lysozyme gene inserted into the T-DNA in Agrobacterium tumefaciens. Two different strategies have been successfully tested to inhibit expression of foreign genes in bacteria using the T4 lysozyme gene as a model system: (a) cloning of the TAC promoter in anti-orientation to the respective gene inside the T-DNA, and (b) insertion of an antisense T4 lysozyme gene under the control of the TAC promoter outside of the T-DNA. Transformation frequencies comparable to controls lacking the T4 lysozyme gene could be achieved when testing potato transformation using both approaches. These strategies might also be useful for the transfer of other genes encoding compounds with detrimental effects to bacteria.