Construction of monopartite geminivirus-based virus-induced gene silencing (VIGS) vectors using a two-component strategy

Virus-induced gene silencing (VIGS) is a rapid and efficient tool to elucidate plant gene functions by inserting a target gene fragment into a viral genome and downregulating the expression of the target gene. Geminiviruses have been considered a promising platform for VIGS, but geminiviral VIGS vectors larger than native viral DNA are often unstable, due to the strict size limitation of their genomes. Here, we present a novel VIGS strategy using two copies of a monopartite geminivirus beet curly top virus (genus Curtovirus) genome that encodes viral genes in the viral and complementary strands. In the first copy, viral strand-encoded genes (V-genes) were replaced by target gene sequences, leaving complementary strand-encoded genes (C-genes) intact, and in the second copy, C-genes were replaced by other target gene sequences, leaving V-genes intact. Co-inoculation of plants with both VIGS vectors supplied all viral proteins necessary for infection, and they systemically spread and induced VIGS. This two-component approach may be used to establish a VIGS system using various species of monopartite geminiviruses.