Post-transcriptional Gene Silencing Using Virus-Induced Gene Silencing to Study Plant Gametogenesis in Tomato.

Loss-of-function analyses are essential to dissect the complex nature of biological processes, including gametogenesis. Virus-induced gene silencing (VIGS) has been widely used in crop species as an amenable and rapid way to generate gene knockdowns. As a transient assay, VIGS circumvents the generation of stable transgenic lines through laborious and time-consuming tissue culture techniques. VIGS involves inoculating plants during early development with genetically manipulated viral constructs carrying an endogenous gene target sequence. The viral infection triggers the host plant gene silencing machinery to process the viral genomic RNA into small RNAs (sRNAs) including the gene complementary region. The sRNAs with complementary sequences to the endogenous gene mediate posttranscriptional gene silencing of the targeted gene. Here, we provide a simple and reproducible VIGS protocol employing the tobacco rattle virus (TRV) in tomato (Solanum lycopersicum cv. M82). As it is stable at later developmental stages this approach is suitable for many traits in tomato including gametogenesis and it can be adapted to other crop species.