Construction Of Expression Vectors For Plant Disease And Stress Resistance Genes

It is an effective way to create new germplasm resources and improve resistance to disease and stress through genetic engineering. Plant expression vector is directly related to the conversion efficiency, genetic stability and expression of the exogenous genes, as well as the safety of genetically modified plants. So the construction of expression vectors which occupies an important position in plant genetic engineering, is the basis of getting useful transgenic plants. In this study, a number of different sources and mechanism resistance genes, such as sprouting resistance genes and different Fusarium graminearum resistance genes were cloned. And they were constructed to wheat and Arabidopsis genetic transformation vectors for the further study about gene function and creating new resistance germplasm. Meanwhile, many different expression vectors were constructed to improve the expression level of foreign gene and achieve higher safety transgenic plants by adding the appropriate elements and constructing the minimal expression cassettes with different promoters. A part of the expression vectors constructed in this study have been used for plant genetic transformation, and plants with phenotypic traits improvement were obtained. The main results are as follows:1.Optimizing the BLF, FUMI, FUMD and LAC gene sequences according to the codon usage bias of Arabidopsis, Triticum aestivum, Hordeumvulgare and Fusarium graminearum; cloning the receptor gene FS1, FS9 and the G protein alpha subunit gene GPA1 from the cDNA of wheat and Arabidopsis; using SOE-PCR technology to amplify a variety of antimicrobial peptides sequences including BLF and MSI99 and antimicrobial peptide-antibody fusion proteins Hvchi-G7, Hvchi-FGMab and UECH42-G7.2. Cloning new tissue-specific expression promoters:amt and lem1; reconstructing the CaMV35S promoter:add intron and Ω sequence to increase the promoter expressional ability and joint it into different expression vectors.3. Cloning different resistance genes and fusion protein into different plant expression vectors.4. Constructing Arabidopsis expression vector containing different detoxification genes:virus-free gene FUMI, FUMD, as well as integration with the antibody H2; the detoxification gene LAC-antibody AFTB1 fusion expression vector; detoxification genes EPO, NHTD expression vectors.5. Cloning a series of antibody, antimicrobial peptides into Arabidopsis transformation vectors, such as A6, B3, D2, E9, H2, G7, F11, ScFvl,20.51, AFTB1, FGMab and PIPP; as well as antibacterial peptides D4E1, MsrAl, AFP2 and MSI99; and fusion proteins D4E1-G7, D4E1-PIPP, D4E1-F11, MsrAl-G7, MsrAl-PIPP, MsrAl-F11, AFP2-ScFv1, AFP2-20.51, AFP2-FGMab, MSI99-ScFvl, MSI99-20.51 and MSI99-FGM Ab.6. Constracting receptor genes FS1 and FS9 fusing with GFP into the plant expression vector to father Arabidopsis transformation.