三七多聚半乳糖醛酸酶抑制蛋白基因的功能分析

doi:10.7668/hbnxb.2018.02.010

摘要/Abstract

摘要： 多聚半乳糖醛酸酶抑制蛋白（PGIP）是植物抗病防御系统的重要组成部分。在前期研究中，从三七中分离得到一个编码PGIP蛋白的基因PnPGIP，PnPGIP在转录水平响应三七根腐病菌茄腐镰刀菌的侵染。为深入分析PnPGIP的功能，构建PnPGIP的原核表达载体，转入大肠杆菌BL21 （DE3）进行大量表达，纯化获得PnPGIP的重组蛋白，并进行体外平板抑菌试验；构建PnPGIP的植物超表达载体，通过根癌农杆菌介导产生了PnPGIP转基因烟草，分析T₂转基因烟草离体叶片对茄腐镰刀菌的抗性。PnPGIP原核重组蛋白对茄腐镰刀菌、胶孢炭疽菌、核盘菌和轮枝状镰刀菌的菌丝生长具有很强的抑制作用，并且随着蛋白量的增加，抑制活性也逐渐增强。qRT-PCR分析结果显示，PnPGIP在T₂转基因烟草中大量表达。此外，与野生型烟草相比，转基因烟草对茄腐镰刀菌的抗性显著增强。PnPGIP是三七中参与茄腐镰刀菌防御反应的重要抗病基因。

关键词: 三七, PGIP, 抗性, 茄腐镰刀菌, 功能验证

Abstract: Polygalacturonase-inhibiting protein(PGIP) plays an important role in plant defense system. PnPGIP, a gene encoding a PGIP protein,was isolated from Panax notoginseng(Burk.) F.H. Chen,and responded the infection of Fusarium solani at the transcriptional level. The objective of this paper was to analyze the function of PnPGIP. The prokaryotic expression vector of PnPGIP was constructed and transferred into Escherichia coli BL21(DE3) for a large amount of expression of the recombinant protein,which was purified to perform the in vitro plate antifungal assay. In addition,the plant overexpression vector was constructed and the transgenic tobacco was obtained by Agrobacterium tumefaciens mediated genetic transformation. The resistance of T₂ transgenic tobacco was analyzed in vivo. The in vitro plate antifungal experiments indicated that the recombinant protein strongly inhibited mycelial growth of Colletotrichum gloeosporioides, Sclerotinia sclerotiorum, F.solani and Fusarium verticillioides, moreover,the inhibitory activity increased with the increased of PnPGIP protein. qRT-PCR analysis showed that PnPGIP was highly expressed in T₂ transgenic tobacco. Furthermore,the resistance of transgenic tobacco to F.solani was significantly enhanced compared with wild type tobacco. PnPGIP is an important disease resistance gene involved in defense response of Panax notoginseng to F.solani infection.

Key words: Panax notoginseng (Burk.) F.H. Chen, PGIP, Resistance, Fusarium solani, Functional verification

中图分类号:

S432
Q786

关瑞攀, 白智伟, 王倩, 唐笔锋, 崔秀明, 刘迪秋. 三七多聚半乳糖醛酸酶抑制蛋白基因的功能分析[J]. 华北农学报, 2018, 33(2): 65-71. doi: 10.7668/hbnxb.2018.02.010.

GUAN Ruipan, BAI Zhiwei, WANG Qian, TANG Bifeng, CUI Xiuming, LIU Diqiu. Functional Analysis of Polygalacturonase-inhibiting Protein Gene from Panax notoginseng[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2018, 33(2): 65-71. doi: 10.7668/hbnxb.2018.02.010.

http://www.hbnxb.net/CN/Y2018/V33/I2/65

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