Abstract
RNA interference (RNAi), a gene-silencing mechanism that involves providing double-stranded RNA molecules that match a specific target gene sequence, is now widely used in functional genetic studies. The potential application of RNAi-mediated control of agricultural insect pests has rapidly become evident. The production of transgenic plants expressing dsRNA molecules that target essential insect genes could provide a means of specific gene silencing in larvae that feed on these plants, resulting in larval phenotypes that range from loss of appetite to death. In this report, we show that the tomato leafminer (Tuta absoluta), a major threat to commercial tomato production, can be targeted by RNAi. We selected two target genes (Vacuolar ATPase-A and Arginine kinase) based on the RNAi response reported for these genes in other pest species. In view of the lack of an artificial diet for T. absoluta, we used two approaches to deliver dsRNA into tomato leaflets. The first approach was based on the uptake of dsRNA by leaflets and the second was based on “in planta-induced transient gene silencing” (PITGS), a well-established method for silencing plant genes, used here for the first time to deliver in planta-transcribed dsRNA to target insect genes. Tuta absoluta larvae that fed on leaves containing dsRNA of the target genes showed an ∼60% reduction in target gene transcript accumulation, an increase in larval mortality and less leaf damage. We then generated transgenic ‘Micro-Tom’ tomato plants that expressed hairpin sequences for both genes and observed a reduction in foliar damage by T. absoluta in these plants. Our results demonstrate the feasibility of RNAi as an alternative method for controlling this critical tomato pest.
Highlights
The mechanism of RNA interference (RNAi), in which small RNAs can rapidly cause post-transcriptional specific gene silencing, has become a powerful tool for analysing gene function in a variety of organisms (Hannon, 2002)
We demonstrated that RNAi for V-ATPase and Arginine kinase (AK), known RNAi target genes among pest insects, significantly reduces target gene expression, increases larval mortality and reduces leaf damage caused by larval herbivory
Both genes studied here have been previously used as RNAi target genes in studies with various agricultural pests, but this is the first report on AK silencing in a lepidopteran species
Summary
The mechanism of RNA interference (RNAi), in which small RNAs can rapidly cause post-transcriptional specific gene silencing, has become a powerful tool for analysing gene function in a variety of organisms (Hannon, 2002). Transgenic plants engineered to express insect dsRNAs emerged as a potential technology after two independent groups proved the concept of applying RNAi to control agricultural insect pests (Baum et al, 2007; Mao et al, 2007). The availability of methods that allow the screening and evaluation of candidate RNAi targets is a critical requisite for developing specific and efficient RNAi-based pest control
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