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作 者 ：黄世容;孟小林;徐进平;鲁伟;王健

期 刊 ：武汉植物学研究 2005年 23卷 4期 页码：376-380

关键词：毕赤酵母；分泌表达；融合蛋白；活性检测；

Keywords:Pichia pastoris, Eliciting expression, Fusion protein, Biological detection,

摘 要 ：将hrpN基因插入毕赤酵母表达载体pPIC6αA的醇氧化酶(AOX1)启动子下游,构建重组表达载体pPIC6αA-hrpN,经电击转化毕赤酵母菌株X-33和杀稻瘟素筛选,得到高效分泌表达harpinEa的毕赤酵母菌株。经初步纯化、SDS-PAGE分析和生物测定,结果显示,诱导96h的此重组菌大量表达harpinEa,表达产物具有诱导烟草超敏感反应的功能,活性高于在大肠杆菌中表达的harpinEa。

Abstract:HrpN gene was placed under the control of AOX1 promoter in a Pichia expression vector pPIC6αA. Through electroporation transformation of X-33 Pichia and Blasticidin selection, a strain of Pichia pastoris capable of secreting harpin_Ea into the medium at high level was obtained. After initially purified, SDS-PAGE analysis of the expression products indicated that harpin_Ea was secreted at high level when the Pichia pastoris expression clone had been induced for 96 hours. The results of bioassay showed that the recombinant harpin_Ea had the ability of eliciting hypersensitive response in tobacco. The activity of this harpin_Ea is higher than that of harpin_Ea expressed in Escherichia coli.

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