Developing a Method to Screen Potential Targets for Artificial RNAi in Aphids
Abstract
Aphids are economically important pest that have developed resistance to many known insecticides. In addition to the direct damage caused by aphid herbivory, aphids are vectors for pathogenic viruses. The goal of this study is to determine if small RNAs can play a role in plant resistance to aphids. RNAi has given rise to a potential new method of insect pest control for crop plants. To further develop understanding of how smRNAs may be used to control insect pests studies were designed to determine candidate genes for targeting using dsRNA in artificial diets and creating amiRNA expressing transgenic Arabidopsis plants. Artificial diets demonstrated that Cathepsin dsRNA administered in complete artificial diet proved more toxic that GFP dsRNA. While transgenic Arabidopsis lines were not tested it was evident that the T-DNA containing the amiRNA was integrated into the plant genome. Further testing will be needed to validate the plants ability to process the amiRNA and effectively target the Cathepsin L gene of the aphid. Transient assays to determine N. benthamiana's ability to process the amiRNA from Agrobacterium infiltration transient assays exhibited mixed results. Both RNA dot blot and RNA gel blot approaches to determining the plants processing ability showed some signs of a mature amiRNA; however, lanes that should have been negative showed hybridization. Also when testing infiltrated tissues with RT-PCR Ct values were much higher than those for housekeeping genes and again negative controls showed positive results.
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- OSU Theses [15752]