植物抗性蛋白“过敏素”的结构与功能关系研究

Bacterial pathogens of both animals and plants use type III secretion machines to secret virulence proteins. The signal that leads to the type III secretion is encoded as stem loop structure in the messenger RNA. Harpin, produced by Erwinia amylovora is secreted via type III secretion machinery. Erwinia carotovora Se9R and Erwinia amylovora belong to the same genus and they use type III secretion machines to secret virulence proteins. We used PCR to amplify hrpN coding region with 75bp upstream segment from pCPP430 which harbors hrpN gene cluster, and cloned it to pGEM-T vector to get pWGF1, which was then chemically transformed into DH5a(pCPP430hrpN－) and electroporated into Se9R. DH5a (pCPP430hrpN－/pWGF1) induced hypersensitive response on tomato leaf and Se9R(pWGF1) showed significantly reduced virulence than Se9R on Chinese cabbage leaf and potato tube. Western-blotting analysis of the CFEP of Se9R (pWGF1) showed production of harpin protein. These results suggest the type III secretion machine in Se9R can recognize secretion signal in the gene of harpin in Erwinia amylovora and secret it as a biologically native form.

梨火疫欧氏杆菌产生的过敏素蛋白可以在其非寄主植物上诱导出对多种病害的抗性。本研究将对过敏素的基因进行缺失、突变和重组，表达变异蛋白，进行个体、细胞和分子水平的植物活性测定，以鉴定出与诱导过敏反应、防卫反应、植物识别等功能相关的结构域。本研究可以为植物抗病基因工程、生防工程菌的构建和新型诱抗剂的设计提供基因材料及理论论据。..