异源Harpin对软腐欧氏杆菌与植物互作关系的影响

THE INFLUENCE OF IN CELL EXPRESSED Harpin_(Ea) ON THE INTERACTION BETWEEN Erwinia carotovora AND HOST AND NON-HOST PLANT

本文通过三亲交配将带有梨火疫欧文氏杆菌功能完整的 hrp基因簇的重组粘粒 p CPP4 30转移到了胡萝卜软腐欧氏杆菌 (Erwinia carotovora) Se9R中。从接合子中提纯的质粒与 p CPP4 30的大小、酶切图谱相同 ;以该质粒为模板 ,用针对 hrp N序列的引物经 PCR扩增得到与 hrp N大小相同的片段 ;用地高辛标记的 hrp N作为探针进行 Southern blotting分析 ,在接合子中的质粒上得到预期的杂交带。接合子可以在烟草叶片上稳定地引起过敏反应。另外 ,接合子可以在寄主马铃薯叶片上引起过敏反应样枯斑 ,表明异源 harpin的表达及功能不受软腐菌的抑制。该接合子在体外产生的果胶酸裂解酶活性与受体菌 Se9R无明显差别。低浓度的接合子在马铃薯块茎上的软腐症状明显低于受体菌 Se9R。外源harpin表达可能不改变软腐欧氏杆菌的致病力、而是通过诱导马铃薯抗性达到减轻发病的作用。

The recombinant cosmid pCPP430 harboring the functional hrp gene cluster of Erwinia amylovora , which produce an elicitor named harpin Ea was transformed into Erwinia carotovora strain Se9R by tri parental mating. Results of restriction enzymatic mapping, PCR amplification with hrpN specific primers and Southern blotting of plasmid DNA with Dig labeled hrpN DNA as probe showed evidents of the successful construction of the conjugant E.carotovora (pCPP430). The conjugant induced hypersensitive response on non host plant tobacco. It also induced hypersensitive like response on leaves of the host plant potato. The conjugant exhibited reduced virulence on potato tube at low inoculum concentration, but retained the same level of pectate lyase activity as that of the recipient Se9R. It was postulated that the apparent reduced virulence of E.carotovora (pCPP430) may be resulted from improved resistance induced by harpin Ea .