The Study Of Cloning Of Npr1 Genes In Two Plants And Transformation Of Poplar

Poplar (Populus) is widely cultivated in the mid-latitude regions of the world. However, diseases of Poplar, especially poplar canker, leaf rust and black leaf spot, seriously affected the growth of poplar, causing large economic losses. SAR (systemic acquired resistance), also is known as broad spectrum resistence. When SAR generated in the plant, the plant could maintain a longer period of increased resistence to multiple pathogens. An important feature of SAR is broad spectrum resistence, SAR induced by one kind of pathogen often show a wide range resistance to viruses, bacteria and fungal pathogen. Thus, it is a great prospect to obtain broad spectrum resistence by regulating SAR response. In recent years, numbers of experiments prove that Arabidopsis thaliana AtNPR1 gene control the SAR response and overexpression AtNPR1 gene can produce SAR response in Arabidopsis while reflect resistance to a wide range of plant pathogens.In this thesis, by using leaves of Poplar(Populus deltoides cv."Zhonglin2001") for exophyte, filter the optimal medium of differentiation of poplar leaves by selecting six different hormone treatment of 6-BA and NAA; AtNPR1 gene is cloned from Arabidopsis and the overexpression vector of pBI121+AtNPR1 is constructed. Then Agrobacterium-mediated genetic transformation of poplar is carried out and Transformed plants are filtered by antibiotic and PCR analysis; at the same time, by the way of the bioinformatics analysis, candidate genes are found in Poplar. And we cloned PtNPR1 gene from Poplar. The main results are summarized as follow:1,Based on the research of directly inducing the adventivebud from Poplar leaves, the best culture medium is MS+ NAA 0.5 mg/L +6-BA 1.0 mg/L +2 % sucrose.2,We cloned gene AtNPR1 from Arabidopsis, constructed the expression vector of pBI121+AtNPR1 which contains CaMV35S constitutive promoter and kanamycin selective gene and the expression vector is introducted into Agrobacterium EHA105 through Freeze Thaw method successfully.3,Through leaf discs transformation system, we transformed over-expression vector pBI121+AtNPR into Poplar Nanlin 95. Five transgenic plants are obtained after kanamycin selecting while three transgenic plants are obtained by PCR analysis.4,By the way of the bioinformatics analysis, 8 candidate genes are found in Poplar which is similar to AtNPR1 gene in Arabidopsis, Further analysis found that one of NPR type gene sequences with the most coincident structure feature, and cloned this gene.In summary, the optimal medium of differentiation of poplar leaves is obtained; AtNPR1 gene is cloned and the overexpression vector of pBI121+AtNPR1 is constructed, genetic transformation of poplar"Nanlin 95"is carried out. At last, three transgenic plants are obtained by PCR analysis; by the way of the bioinformatics analysis, NPR type genes are found in the poplar and the PtNPR1 gene is cloned. It is good for cultivating species with high-yield, quality and broad spectrum resistence. It provided materials for further research on resistance mechanism, and also laid a theoretical foundation for resistance genetic engineering of other tree plants.