Virus-induced gene silencing in various Prunus species with the Apple latent spherical virus vector

Abstract

Virus-induced gene silencing (VIGS) has been used as a rapid and effective tool for functional analysis of genes in various plants, including woody fruit tree species. We previously reported the successful induction of VIGS of the endogenous PHYTOENE DESATURASE (PDS) gene in apricot (Prunus armeniaca L.) using Apple latent spherical virus (ALSV) vectors. In contrast, our attempts to infect Japanese apricot (Prunus mume Siebold & Zucc.) with ALSV vectors was unsuccessful, suggesting that species- and/or cultivar-dependent differences of ALSV susceptibility may exist in Prunus. In this study, we investigated whether this VIGS-based gene evaluation system using ALSV vectors was applicable to seven Prunus species, including apricot, sweet cherry (Prunus avium L.), almond [Prunus dulcis (Mill.) D. A. Webb.], peach (Prunus persica Batsch), Japanese apricot, Japanese plum (Prunus salicina Lindl.), and European plum (Prunus domestica L.). ALSV vectors carrying part of the apricot PDS gene sequence were amplified in Nicotiana benthamiana, and inoculated into the cotyledons of Prunus seedlings by particle bombardment. Typical PDS-silenced phenotypes, characterized by uniform discoloration of the upper leaves, were observed in sweet cherry and some cultivars of apricot and almond several weeks after inoculation. The amounts of PDS mRNA in the infected leaves were significantly reduced, while a 21nt antisense small RNA, which was assumed to play a central role as a guide RNA in PDS mRNA degradation, was highly accumulated. However, ALSV infection of Japanese apricot, Japanese plum, European plum, and the other cultivars of apricot and almond was unsuccessful. Furthermore, although the infection rate of ALSV in peach was high, severe pale spots (a viral infection symptom) were observed in the infected leaves. These results collectively suggested that the efficiency of ALSV infection and VIGS could vary depending on species and/or cultivar in Prunus. The possible use of the ALSV-mediated VIGS system for functional analysis of genes in Prunus is discussed.