Molecular cloning and RNA interference of Cullin-1 gene in the codling moth Cydia pomonella

Monday, November 17, 2014: 8:24 AM
A106 (Oregon Convention Center)
Stephen Ireland , Molecular, Cellular and Developmental Biology, University of Michigan, Ann Arbor, MI
Jinda Wang , Department of Entomology, Nanjing Agricultural University, Nanjing, China
Douglas Knipple , Entomology, Cornell University, Geneva, NY
RNAi-based pest management practices have the potential to be an alternative to, augment or replace existing molecular based strategies. However, some insect taxa are less susceptible to the induction of RNAi than others, such as the lepidoptera. Here we describe experiments to investigate the induction of RNAi in the codling moth Cydia pomonella, a major lepidopteran pest of apple, pear and walnut. Prior to a knockdown screen, we demonstrated that fluorescently labelled siGFP and dsGFP RNAs were transmissible through the gut epithelium. We then synthesized double-stranded RNAs (dsRNAs) targeting five C. pomonella genes orthologous to those that have well defined deleterious phenotypes in Drosophila melanogaster. The dsRNAs were administered to neonate larvae by incorporation into the growth media. None of the dsRNA target treatments affected larval viability in vitro, however dsRNAs induced a visible phenotype in response to Cpcul1-dsRNA. A significant stunting of growth was observed. A dose-dependent RNAi effect was observed as relative expression of Cpcul1 showed a decrease consistent with an increasing dose of Cpcul1-dsRNA. However, despite their reduced size, Cpcul1-dsRNA-treated larvae molted normally and matured to adulthood in a manner similar to controls. Additionally, the stunting effect induced by Cpcul1-siRNA was found to be similar to that induced by Cpcul1-dsRNA.