利用酵母双杂交系统筛选感病番茄中蔬四号与无毒蛋白AvrPto、AvrPtoB互作的蛋白

doi:10.3864/j.issn.0578-1752.2011.23.022

摘要/Abstract

摘要： 【目的】利用酵母双杂交系统在感病番茄中蔬四号中筛选分别与AvrPto、AvrPtoB互作的蛋白，研究AvrPto、AvrPtoB作为毒性因子在感病番茄中的作用通路。【方法】提取感病番茄中蔬四号的总RNA，合成中蔬四号cDNA文库，分别构建AvrPto和AvrPtoB的诱饵载体，从中蔬四号cDNA文库中筛选分别与AvrPto、AvrPtoB互作的蛋白，并对阳性克隆进行分析和鉴定。【结果】最终筛选到7个与AvrPto互作的蛋白，2个与AvrPtoB互作的蛋白，核酮糖-1,5-二磷酸（RuBP）羧化酶/加氧酶小亚基和AvrPto、AvrPtoB都能够互作，说明AvrPto、AvrPtoB参与破坏植物的光合作用；另外与AvrPto互作的蛋白质还包括叶绿体原卟啉原氧化酶、叶绿体醛缩酶、叶绿体延伸因子TufA、翻译延伸因子等参与植物光合作用蛋白。【结论】分析筛选到的蛋白主要是参与植物光合作用的蛋白，推测AvrPto、AvrPtoB通过与植物光合作用有关的蛋白互作，或者影响卡尔文循环，或者影响叶绿素和相关蛋白质的合成，从而破坏植物光合作用，促进侵袭位点叶肉细胞衰老、死亡，进而引起坏死症状。

关键词: 番茄细菌性斑点病菌, 无毒基因, 酵母双杂交

Abstract: 【Objective】 The objective of this study is to research the function of AvrPto and AvrPtoB as virulence factor in the susceptible tomato by yeast two-hybrid system. 【Method】 The total RNA of Zhongshusihao was extracted, and dscDNA was amplified by long distance PCR. pGBKT7-AvrPto and pGBKT7-AvrPtoB were constructed. The AvrPto and AvrPtoB interactive proteins were screened by two-hybrid system. 【Result】 In the susceptible tomato cultivar Zhongshusihao cDNA library, 7 proteins interacting with AvrPto were identified, 2 proteins interacting with AvrPtoB were identified. Ribulose bisphosphate carboxylase small chain which can interact with both AvrPto and AvrPtoB showed that AvrPto and AvrPtoB may participate in destroying host photosynthesis. Other proteins such as chloroplastic protoporphyrinogen IX oxidase, chloroplast aldolase which participate in host photosynthesis, and chloroplast elongation factor TufA, transcription elongation factor were also participated in protein synthesis. 【Conclusion】According to the results, it was presumed that AvrPto, AvrPtoB interacting with photosynthesis proteins affected Calvin cycle, the synthesis of chlorophyl related protein. By this way, mesophyll cell of host invation site died.

Key words: Pseudomonas syringae pv. tomato, avirulence gene, yeast two-hybrid system

张月娟, 赵廷昌, 杨玉文. 利用酵母双杂交系统筛选感病番茄中蔬四号与无毒蛋白AvrPto、AvrPtoB互作的蛋白[J]. 中国农业科学, 2011, 44(23): 4939-4944.

ZHANG Yue-Juan, ZHAO Ting-Chang, YANG Yu-Wen. Screening of the Interactive Proteins of Avirulent Proteins AvrPto or AvrPtoB in Susceptible Tomato Cultivar Zhongshusihao by Yeast Two-Hybrid System [J]. Scientia Agricultura Sinica, 2011, 44(23): 4939-4944.

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