Abstract:

Genetic transformation with multiple genes is one of the popular researches in gene engineering. Difensin which is resistant to many kinds of virus, bacterium and fungal, and Gastrodia Antifungal Protein (GAFP) which has the resistant ability to more than twenty kinds of fungal were used for the vector construction. Also, the mechanisms of anti-disease by two genes are different from each other. By use of DNA recombination method a vector (pBin35SGAFP- NP1) that contains the two anti-disease genes: NP1 and GAFP was constructed. Mediated by Agrobacterium tumefaciens, the two disease-resistance genes were transferred into tobacco using leaf-disk method. Results of PCR and PCR-Southern determination indicated that NP1 and GAFP genes were integrated into the tobacco genome. In vitro testing of transgenic plants to virus, bacterium and fungal showed that they had the resistance to the growth of Agrobacterium tumefaciens, Trichoderma reesei, Sphaeropsis sapinea, and it was appeared that NP1 and GAFP genes were expressed efficiently in transgenic tobacco and the transgenic plants had disease-resistant ability to pathogens. Therefore, by using this vector transgenic plants with the two anti-disease genes were obtained which were proved to be expressed efficiently and the ability of anti-disease could be increased.

Key words: NP1, Gastrodia Antifungal Protein (GAFP), vector with two anti-disease genes, transgenic tobacco, disease-resistance