Development of an in vitro RNA silencing method for the functional analysis of genes affecting early tomato fruit growth

RNA silencing (RNAi) is a genetic regulatory process mediated by dsRNA. The expression of gene constructs containing inverted repeats (hpGenes) elicits RNAi either transiently or permanently, depending on their stable integration into the genome. The stable introduction of a hpGene into the genome is a useful tool for studying gene function, however it is a lengthy process. To reduce the length of gene functional analysis, methods for transient RNAi in tomato fruits have been developed. Although effective, these approaches present some disadvantages (e.g. they can cause lesions to the fruit itself). We propose a novel method based on in vitro transient RNAi specifically designed to study genes involved in early stages of tomato ovary/fruit development. The method uses pre-anthesis flower buds collected from parthenocarpic auxin-synthesising tomatoes which are able to substain in vitro ovary/fruit growth without phytohormones. Young parthenocarpic flower buds are inoculated with agrobacteria bearing hpGenes using a procedure that avoids damages to the ovary.