|WANG, JINDA - Cornell University
|GU, LIUQI - Cornell University
|IRELAND, STEPHEN - Cornell University
|KNIPPLE, DOUGLAS - Cornell University
Submitted to: Gene
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/2/2015
Publication Date: 11/10/2015
Publication URL: http://handle.nal.usda.gov/10113/62786
Citation: Wang, J., Gu, L., Ireland, S., Garczynski, S.F., Knipple, D.C. 2015. Phenotypic screen for RNAi effects in the codling moth Cydia pomonella. Gene. 572:184-190.
Interpretive Summary: Codling moth is a key insect pest of apple, pear and walnut, and new methods that are safe and effective are needed to control it. Scientists at the USDA-ARS, Yakima Agricultural Laboratory in Wapato, WA and at Cornell University, New York State Agricultural Experiment Station, Geneva, NY developed and demonstrated an RNA interference technique that is effective for altering gene expression in codling moth. They introduced RNA molecules into codling moth larvae that inhibited their growth by reducing the expression of the specific protein called Cullin-1. These results provide the methods and show the potential for the development of RNA interference as a technique to reduce codling moth development, survival, and reproduction in orchards.
Technical Abstract: RNAi-based technologies have the potential to augment, or replace existing pest management strategies. However, some insect taxa are less susceptible to the induction of the post-transcriptional gene silencing effect 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, fluorescently labeled small interfering RNA (siRNA) and double-stranded RNA (dsRNA) derived from green fluorescent protein (GFP) coding sequence were shown to be transmissible through the gut epithelium. Next, dsRNAs were made for C. pomonella genes orthologous to those that have well defined deleterious phenotypes in Drosophila melanogaster. A screen was conducted using dsRNAs encoding cullin-1 (Cpcul1), maleless (Cpmle), musashi (Cpmsi), a homeobox gene (CpHbx), and pumilio (Cppum). The dsRNAs designed from these target genes were administered to neonate larvae by incorporation into the growth medium. None of the dsRNA target treatments affected larval viability, however Cpcul1-dsRNA induced a significant stunting of growth, with the average length of larvae about 3 mm, compared to about 4 mm in the control groups. Measurement of Cpcul1 transcript levels by quantitative real-time PCR (qRT-PCR) revealed a dose-dependent RNAi effect in response to increasing amount of Cpcul1-dsRNA. Despite their reduced size, Cpcul1-dsRNA-treated larvae molted normally and matured to adulthood in a manner similar to controls. In an additional experiment, Cpcul1-siRNA was found to induce similar stunting effect as that induced by Cpcul1-dsRNA.