A computational approach to execute siRNA generating hotspots targeting dual DNA and RNA viral infections in potato

Among biotic stresses afflicting potato plants, viruses are the most damaging and are responsible for large economic losses worldwide. Co-infections with multiple viruses are common in potato, with an enhanced disease impact being observed in affected plants. RNA interference (RNAi) provides an applied methodology to selectively reduce the expression of targeted genes throughthe expression of sequence-specificshort interfering RNAs (siRNAs). This silencing mechanism can be implemented to induce resistance against multiple viruses in transgenic plants through the endogenousdelivery of siRNA cassettes. The current study was aimed to identify the efficient siRNA execution sites in dominating viral genomes to simultaneously target both DNA and RNA viruses in potato. To achieve this objective, we followed a computational approach to identify the viral silencing targets by comparative pairwise sequence analysis of different isolates of Potato leafrollvirus(PLRV; + single-stranded(ss) RNA virus) and Tomato leaf curl New Delhi virus(ToLCNDV; ssDNA virus). The identified consensus sequences [300bp of PLRV-coat protein (CP); 180bp of ToLCNDV-precoat protein (AV2)]were further used as template sequences to predict thelikely siRNAs execution sites and to characterizetheir putative thermodynamic attributes. The identified template sequences were computationally tested for triggering a siRNA-mediated targeting of viral genomes and proved to be highly efficient and site-specific. This methodology could be applied forengineering anRNAi-mediatedvirus resistance in transgenic plants with commercial applications.