Applications of biotechnology for crop enhancement in disease resistance and nutrition

Developments in plant biotechnology are providing solutions for addressing the problems and challenges of sustainable agriculture and providing healthy food. Engineering disease resistant and nutritionally improved agricultural crops is integral to the application of plant biotechnology. In this thesis study, our core interests are on (i), studying and manipulating the economically important plant disease Fusarium head blight (FHB) by generating CRISPR-edited Arabidopsis resistant mutant plants; and (ii), producing nutritionally enhanced tomato plants with a two-in-one strategy. FHB caused by Fusarium graminearium (Fg) is a devastating disease of crops especially wheat and barley, resulting in significant yield loss and reduced grain quality. Fg also produces mycotoxins, which are chemicals that are toxic to humans and livestock upon consumption. In order to examine potential susceptibility genes in barley and to enhance host resistance to FHB, two susceptibility genes, 2-oxoglutarate Fe(II)-dependent oxygenase (2OGO) and ethylene insensitive 2 (EIN2) genes were chosen for this study. Mutation of the 2OGO gene has been shown to enhance plant defense genes’ expression, leading to FHB resistance in Arabidopsis, indicating that 2OGO is a plant immunity suppressor. Previous reports showed that both loss-of-function Arabidopsis EIN2 gene and attenuated expression of wheat EIN2 gene conferred resistance to Fg, leading to the speculation that the ethylene signaling pathway is hijacked by Fg. We have used CRISPR/Cas9-gene editing to precisely mutate these two Arabidopsis FHB susceptibility genes. Our results indicated that both At2OGO- and AtEIN2-gene knocked-out (KO) mutants were resistant to FHB such that Fg growth was greatly impeded. A critical part of this study is that phenotype could be restored in gene complementation assays using the barley gene orthologues Hv2OGO and HvEIN2 in the At2OGO- and AtEIN2-KO mutants, thus demonstrating their involvement in FHB susceptibility, and indicating the molecular targets for similar gene editing of these genes in barley plants. Genistein is one of the major isoflavones found abundantly in soybeans. It is a promising medicinal compound with anti-inflammatory, anticarcinogenic, anti-Alzheimer’s and anti-osteoporosis effect due to its phytoestrogenic and antioxidant properties. Since soy products are not commonly consumed in the western diet as compared to the eastern diet, genetically engineered Moneymaker tomato which produces genistein provides a natural route to increase the intake of genistein in food without relying on supplements. We have successfully engineered Moneymaker tomato to produce a level of genistein, that is up to 250-fold the levels found in the wildtype (WT) tomato plants. Transgenic tomatoes expressing genistein, were used to study the compound’s anti-Alzheimer’s effects on the amyloid β-expressing C. elegans disease model. Our results show that a combination of feeding genistein and lycopene can significantly delay Alzheimer’s disease progression and reduce neuronal damage.Ph.D.Includes bibliographical reference