Cloning Of Resistance-Related Genes Against Fusarium Head Blight And Construction Of Expression Vectors For Monocotyledonous Plants

Fusarium head blight caused by species of Fusarium is an economically important disease on grain cereal crops, which not only can cause grain shrinkage and yield loss, but also produce a variety of mycotoxins that can potentially pose serious threat on human health and animal production. However, because of limited amount of available germ plasm with natural resistance to fungal diseases?the complex of the inheritable mech-anisms of resistance and limitation of the identification of resistance, the enhancement of scab resistance in crop plants has been a major chanllenge. So, Plant genetic engineering provides the new approach to increase crop resistance by transferring the resistance genes into crop plants. In this research, the different resistance genes were cloned and different expression vectors were constructed for plant genetic transformation. The major results were followed:1. We chose the preferential codon of Triticum aestivum, designed and synthesized oligo-nucleotide, then we obtained ZmGC and ECH42 by SOE-PCR. The fusion gene (OAPAR) of mutiple genes mediated with Lp4-2A was constructed, and the gene with different signal peptide was secreted different cell position. In addition, using (G4S)3 as linker, the AMP-scFv fusion genes was obtained by SOE-PCR.2. Lemmas-specific expression vector for Monocotyledonous plants was constructed. The crop plants transformed with Lem2 promoter/resistant gene construction will display cell-and development-specific expression of resistance gene. Then the ability against pathogen invasion of transgenic crops will increase and the negative influence of constitutive expression will be avoided.3. A binary plasmid (PTR-LYB-AG) containing the selectable marker gene and the gene of interest in separate T-DNAs was constructed. Taking advantage of Agrobacteri-um mediated transfer, the primary transformant in which the selectable marker gene and the gene of interest subsequent integrate at sufficiently unlinked sites will be generated. Following by genetic out segregation in the progenies, we will screen the marker-free transgenic plants. 4. Different resistance related genes against Fusarium head blight were cloned to different expression vectors for blolistic or A. tumefaciens-mediated transformation. And expression vectors for A. tumefaciens-mediated transformation were transferred into A. tumefaciens strain GV3101.5. Surface antigen hemagglutinin (HA) and neuraminidase (NA) genes?matrix protein 1 (M1) and matrix protein 2 (M2) genes of H5N1 subtype of an avian influenza virus from China were cloned into endosperm-specific expression vectors (pLYA and pBAR-LYA). Now the genes were transferred into wheat by blolistic transformation method.