Double-Strand RNA (dsRNA) Delivery Methods in Insects: Diaphorina citri

RNAi is a gene-silencing mechanism conserved in the vast majority of eukaryotes. It is widely used to study gene function in animals due to the ease of eliciting gene knockdown. Beyond research applications, RNAi technology based on exogenous dsRNA is a promising candidate for next generation insect pest control. An advantage of using RNAi is that design of dsRNA essentially requires only the sequence of the target gene. The greatest challenge, however, is dsRNA delivery for large-scale insect control. Delivery methods that have widely been used are oral, injection, or via soaking. Unfortunately, each insect presents its own challenges owing to the differences in the presence of dsRNA degrading enzymes, cellular uptake efficiency, expression of core RNAi machinery, the nature of the target gene, the concentration and persistence of the dsRNA, as well as the particular way of feeding of each insect, which together cause variations in the efficiency of RNAi. In this chapter, a protocol for the synthetic production of dsRNA is described along with three methods for delivery that have been successful in one of the more problematic insects, Diaphorina citri.