Engineering broad-spectrum resistance against RNA viruses in potato

Abstract

RNA silencing technology has become the tool of choice for inducing resistance against viruses in plants. A significant discovery of this technology is that double-stranded RNA (dsRNA), which is diced into small interfering RNAs (siRNAs), is a potent trigger for RNA silencing. By exploiting this phenomenon in transgenic plants, it is possible to confer high level of virus resistance by specific targeting of cognate viral RNA. In order to maximize the efficiency and versatility of the vector-based siRNA approach, we have constructed a chimeric expression vector containing three partial gene sequences derived from the ORF2 gene of Potato virus X, Helper Component Protease gene of Potato virus Y and Coat protein gene of Potato leaf roll virus. Solanum tuberosum cv. Desiree and Kuroda were transformed with this chimeric gene cassette via Agrobacterium tumefaciens-mediated transformation and transgenic status was confirmed by PCR, Southern and double antibody sandwich ELISA detection. Due to simultaneous RNA silencing, as demonstrated by accumulation of specific siRNAs, the expression of partial triple-gene sequence cassette depicted 20% of the transgenic plants are immune against all three viruses. Thus, expression of a single transgene construct can effectively confer resistance to multiple viruses in transgenic plants.