Efficient Protein Expression and Virus-Induced Gene Silencing in Plants Using a Crinivirus-Derived Vector

Abstract

Plant virus-based vectors are valuable tools for recombinant gene expression and functional genomics for both basic and applied research. In this study, <i>Lettuce infectious yellows virus</i> (LIYV) of the genus <i>Crinivirus</i> was engineered into a virus vector that is applicable for efficient protein expression and virus-induced gene silencing (VIGS) in plants. We examined gene replacement and &ldquo;add a gene&rdquo; strategies to develop LIYV-derived vectors for transient expression of the green fluorescent protein (GFP) reporter in <i>Nicotiana benthamiana</i> plants. The latter yielded higher GFP expression and was further examined by testing the effects of heterologous controller elements (CEs). A series of five vector constructs with progressively extended LIYV CP sgRNA CEs were tested, the longest CE gave the highest GFP expression but lower virus accumulation. The whitefly transmissibility of the optimized vector construct to other host plants, and the capability to accommodate and express a larger gene, a 1.8 kb &beta;-glucuronidase (GUS) gene, were confirmed. Furthermore, the LIYV vector was also validated VIGS by silencing the endogenous gene, <i>phytoene desaturase</i> (PDS) in <i>N. benthamiana</i> plants, and the transgene GFP in <i>N. benthamiana</i> line 16c plants. Therefore, LIYV-derived vectors could provide a technical reference for developing vectors of other economically important criniviruses.