Studies On Transgenic Wheat With Preharvest Sprouting Resistant Genes And Fusarium Head Blight Resistant Genes

Preharvest sprouting (PHS) in wheat is defined as the germination of grains in the ears, usually induced by wet conditions after maturity and before harvest. PHS not only reduced crop quality and production, but also casused great economic loss and crop supplement threat. PHS is a world wide disaster in wheat production, which emerged in many wheat growing countries. In China, the vast majority of areas for wheat production, accounting for 83% of acreages in the country, are affected by PHS. Wheat germplasm with resistance to PHS is limited. Therefore, development of new strategies through transgenic technology provides alternatives for the improvement of PHS resistance in wheat.Fusarium head blight (FHB), caused by Fusarium graminearum, is a destructive disease of wheat in warm and humid regions around the world. Although FHB has been the focus of much research, this disease continues to cause serious yield losses and to threaten human and animal health via the contamination of food and feed with Fusarium mycotoxins. FHB is one of most important diseases frequently happens in the middle and lower regions of the Yangtze River. However, traditional wheat breeding programmers have been limited by lacking of effective germplasm. Therefore, novel sources of FHB resistance are required, and genetic engineering provides an additional approach to increase FHB resistance in wheat.In this study, PHS resistant genes were introducted into wheat cultivar Zhengmai9023, and FHB resistant genes were integrated into wheat cultivar Yangmai158. We also co-transformed these two kinds of resistance-related genes into wheat cultivar Zhengmai9023. We had obtained transgenic wheat plants containing these genes. The main results are as follows:1.Three PHS resistant expression cassettes including maize vpl coding sequence and different kinds of promoters (Dvp1-intron-vp1-nos, vp1-UTvp1-nos, Dvp1-vp1-nos) were transformed into an elite wheat cultivar Zhengmai9023 via particle bombardment, respectively. To transgenic plants were selected in the presence of Bialaphos and identified by PCR. Then we obtained ten transgenic plants. Seven of them contained Dvp1-intron-vp1-nos, two contained vp1-UTvp1-nos and one contained Dvp1-vp1-nos.2. The wheat cultivar Yangmai158 was used for co-bombardment with three anti-fungal peptide expression cassettes:BLF, LfcinB, BLF-A6. Transgenic plants were selected on media supplemented with Bialaphos. PCR identification results showed that there was one transgenic wheat plant containing BLF-A6 in To generation.3.Using Bar gene as a selectable maker, we transformed three PHS resistant genes (vpl, Basi-Basi, AsGlo1-Basi) and one antifungal gene (AFP2) into wheat cultivar Zhengmai9023. PCR identification results indicated that there were eight transgenic wheat plants in To generation. These transgenic wheat plants contained all five alien genes. We further studied the inheritance and integration pattern of eight transgenic wheat plants by PCR assay and Southern blotting analysis. Furthermore, we performed RT-PCR assay to detect the mRNA expression of target genes in transgenic wheat lines. Then seed GI and spike PS, DS were investigated to assay seed dormancy of transgenic wheat lines. Also, we assessed the FHB resistance of transgenic wheat lines. In summary, the transgenic wheat lines can be divided into three groups. In the first group, the integration and expression of PHS resistant genes and AFP2 gene significantly improved the PHS resistance and FHB resistance of transgenic wheat lines. Transgenic wheat lines in the second group contained or expressed PHS resistant genes, and their PHS tolerance was significantly increased. While in the third group, the percentage of infected spikelets of transgenic wheat lines was significantly reduced because of the integration and expression of AFP2 gene.