RNA interference-based resistance against a legume mastrevirus

Abstract

<p>Abstract</p> <p>Background</p> <p>RNA interference (RNAi) is a homology-dependant gene silencing mechanism and has been widely used to engineer resistance in plants against RNA viruses. However, its usefulness in delivering resistance against plant DNA viruses belonging to family <it>Geminiviridae </it>is still being debated. Although the RNAi approach has been shown, using a transient assay, to be useful in countering monocotyledonous plant-infecting geminiviruses of the genus <it>Mastrevirus</it>, it has yet to be investigated as a means of delivering resistance to dicot-infecting mastreviruses. <it>Chickpea chlorotic dwarf Pakistan virus </it>(CpCDPKV) is a legume-infecting mastrevirus that affects chickpea and other leguminous crops in Pakistan.</p> <p>Results</p> <p>Here a hairpin (hp)RNAi construct containing sequences encompassing part of replication-associated protein gene, intergenic region and part of the movement protein gene of CpCDPKV under the control of the <it>Cauliflower mosaic virus </it>35S promoter has been produced and stably transformed into <it>Nicotiana benthamiana</it>. Plants harboring the hairpin construct were challenged with CpCDPKV. All non-transgenic <it>N. benthamiana </it>plants developed symptoms of CpCDPKV infection within two weeks post-inoculation. In contrast, none of the inoculated transgenic plants showed symptoms of infection and no viral DNA could be detected by Southern hybridization. A real-time quantitative PCR analysis identified very low-level accumulation of viral DNA in the inoculated transgenic plants.</p> <p>Conclusions</p> <p>The results presented show that the RNAi-based resistance strategy is useful in protecting plants from a dicot-infecting mastrevirus. The very low levels of virus detected in plant tissue of transgenic plants distal to the inoculation site suggest that virus movement and/or viral replication was impaired leading to plants that showed no discernible signs of virus infection.</p