Plant improvement for insect resistance: Testing of the candidate organism Beauveria bassiana, transgenic tobacco expressing protease inhibitors, and rapid screen of insect resistance genes in an agroinfiltration transient expression system
Date of Award
Doctor of Philosophy
Plants, Soils, and Insects
C. Neal Stewart, Jr
Feng Chen, Ranjan Ganguly, Bonnie Ownley
This study focused on three aspects of plant improvement for insect resistance including: testing of candidate organisms for their production of insecticidal proteins, testing of transgenic plants expressing insect resistance genes, and testing novel systems for the evaluation of insect resistance genes. In the initial part of this study, the candidate fungus Beauveria bassiana was tested for its production of insecticidal proteins through a series of insect bioassays containing fungal protein extracts. These extracts were shown to be orally toxic to Plutella xylostella (diamondback moth) and Spodoptera frugiperda (fall armyworm). Assays involving protease treatments significantly decreased mortality indicating the presence of a protein based oral toxin. The following research tested transgenic tobacco plants expressing proteinase inhibitors from Brassica oleracea (cabbage) and Manduca sexta (tobacco hornworm) on the insect pests Helicoverpa zea (corn earworm) and Heliothis virescens (tobacco budworm). Insects fed transgenic tobacco were able to adapt to the recombinant proteinase inhibitors to varying degrees and resulted in no major impacts on insect growth and development. The last part of this study tested a novel insect resistance gene screening system. Agroinfiltrated tobacco transiently co-expressing genes encoding GFP with either a known insecticidal protein (Bt Cry1Ac) or a candidate gene (Brassica oleracea proteinase inhibitor, BoPI) were fed to larval H. zea. Insects fed the known insecticidal protein experienced high mortality. Insects fed tobacco expressing GFP and BoPI showed significant decreases in growth compared to those fed GFP only tissue. Insects feeding on GFP only tissue showed unexpected increases in growth and development compared to insects fed control tissue. Agroinfiltration coupled with an insect bioassay constitutes an efficient system for the evaluation of candidate insect resistance genes.
Leckie, Brian M., "Plant improvement for insect resistance: Testing of the candidate organism Beauveria bassiana, transgenic tobacco expressing protease inhibitors, and rapid screen of insect resistance genes in an agroinfiltration transient expression system. " PhD diss., University of Tennessee, 2008.