Production of Potato Resistant Plant to PVX using an RNA Silencing Mechanism

Viruses are the main causes of potato yield loss. Potato X Potexvirus (PVX) is one of the most important potato viruses and the use of resistant varieties is the principal way to control it. In this study, we tried to produce potato plants resistant to this virus using an RNA silencing technique. To this end, a hairpin construct of P25 of PVX was made in pFGC5941 under control of the CaMV 35S promoter with two consecutive cloning steps. The P25 protein is a suppressor of RNA Silencing in viral host plants. The recombinant vector was transferred into Agrobacterium tumefaciens LBA4404 strain. Leaf and internode pieces of potato (Agria variety) were transformed by Agrobacterium. Following induction of callus, shoot and root, regenerated transgenic plants were selected and micro-propagated. Molecular analysis by PCR on DNA of plants and by RT-PCR on their RNA confirmed the presence and the expression of the transgene. Transgenic plants were propagated and then inoculated mechanically by PVX in the greenhouse. ELISA results using PVX-coat protein antibody showed resistance of two transgenic lines to PVX. Molecular confirmation of these potato lines was performed using PVX coat protein primers by RTPCR on plant RNAs. The results of this research have led to the production of two independent lines of potato that are resistant to PVX and future work will involve analysis of resistance in field conditions and the production of their mini-tuber.