Evaluation of methods of dsRNA delivery for RNA Interference as a functional tool in the Colorado potato beetle, Leptinotarsa decemlineata (Say)

RNA interference is used as a tool for the functional analysis of genes.  Introducing double-stranded RNA into an organism suppresses expression of the corresponding gene.  Depending on the expression pattern of the target gene of interest, effective functional analysis may demand RNAi that is systemic and/or persistent.  We are investigating different methods of dsRNA delivery as a tool for the functional analysis of genes for the Colorado potato beetle, Leptinotarsa decemlineata (Say). L. decemlineata is a devastating pest to the plant family Solanaceae and therefore an important model for the study of insect-plant interactions.  As a proof of concept, we are focusing on the laccase-2 gene.  Silencing laccase-2 has been shown to result in the reduction of cuticular pigmentation in the western corn rootworm, Diabrotica virgifera virgifera (LeConte) and the red flour beetle Tribolium castaneum.  The conspicuous but not immediately lethal phenotypic effect of suppressing laccase-2 by RNAi makes it an ideal target gene for investigating RNAi as a functional tool.  The RNAi-induced knockdown effect of laccase-2 can be evaluated phenotypically and by qPCR.  Here we use RT-qPCR to evaluate functional RNAi in the egg and larval stages of L. decemlineata following different methods of dsRNA delivery.