Submitted to: National Fusarium Head Blight Forum Proceedings
Publication Type: Proceedings
Publication Acceptance Date: November 13, 2002
Publication Date: December 7, 2002
Citation: Somleva, M., Okubara, P.A., Blechl, A.E. 2002. Transgene expression in spring wheat (triticum aestivum l.) transformed with candidate anti-fusarium genes. National Fusarium Head Blight Forum Proceedings. 42-45 Interpretive Summary: Fusarium Head Blight (FHB) or scab is a disease of wheat and barley that has caused billion dollar losses to farmers and processors in the past decade. The disease is caused by a fungus that invades flowers as they open. There are few sources of natural genetic resistance to FHB among cultivated wheats. To supplement and extend these partial resistances, we have transformed wheat with genes that code for proteins designed to fight Fusarium invasion. The proteins include enzymes that degrade fungal cell walls. Measurements of the levels of RNA produced by our transgenes show a range of values, from very high to barely detectable. We developed a new method that allows us to visualize the location of these RNAs in the tissues of the plant. Results using this method show that our transgenes are expressed in young flowers, in the tissues most susceptible to fungal invasion. Thus, our anti-fungal proteins will be made in the right tissues at the right stage of development to be effective.
Technical Abstract: Host plant resistance is the most efficient and cost-effective way to protect the wheat crop from Fusarium Head Blight (FHB). Our aim is to create new germplasm sources of Fusarium resistance by using genetic transformation to introduce novel anti-Fusarium (AF) genes into wheat. We have designed and constructed a set of transformation vectors that fuse the maize Ubi1 promoter to AF genes that target either the Fusarium cell walls or membranes or that mitigate the cellular toxicity of the mycotoxins synthesized by the fungus during infection. Five of our genes come from Fusarium, including coding regions for a glucanase, exochitinase, endochitinase, DON acetyltransferase and a trichothecene efflux pump. The 5' untranslated regions of these genes have been modified to more closely resemble those of well-translated wheat genes. We have obtained a number of transformants carrying these genes in the cultivar Bobwhite, a hard white spring wheat with good tissue culture properties and moderate Type II resistance. We use semi-quantitative RT-PCR to measure expression levels of our transgenes in various tissues of plants of early and later generations. We have found transformants with high levels of expression for the two Fusarium-derived chitinase genes, as well as for a gene from wheat that encodes a thaumatin-like protein, but not for the Fusarium-derived glucanase or DON acetyltransferase genes. In general, expression levels are higher in later generations than in earlier ones. Our highest expressor plants have been or are being tested for Type II resistance. In order to more precisely localize transgene expression, we have developed an RT-PCR method for in situ detection of transcripts in whole mount sections of wheat tissues and organs. This method is being refined to better visualize expression in young florets.