酵母双杂交筛选灰飞虱体内水稻黑条矮缩病毒 p10 的互作蛋白

doi:10.3969/j.issn.1001-7216.2013.06.010

中国水稻科学 ›› 2013, Vol. 27 ›› Issue (6): 633-638.DOI: 10.3969/j.issn.1001-7216.2013.06.010

酵母双杂交筛选灰飞虱体内水稻黑条矮缩病毒 p10 的互作蛋白

徐秋芳,陈晴晴,张金凤,李硕,倪海平,周益军^*

江苏农业科学院植物保护研究所/江苏省植物病毒病诊断检测中心, 南京 210014；

收稿日期:2013-04-27 修回日期:2013-05-17 出版日期:2013-11-10 发布日期:2013-11-10
通讯作者: 周益军*
基金资助:
国家自然科学基金资助项目（31000841）；公益性行业（农业）科研专项（201003031）；江苏省农业科学技术自主创新资金项目\[CX（13）5019\]。

Screening of RBSDV p10 Interacting Proteins in Small Brown Planthoppers by Yeast Two Hybrid System

XU Qiufang, CHEN Qingqing, ZHANG Jinfeng, LI Shuo, NI Haiping, ZHOU Yijun^*

Institute of Plant Protection, Jiangsu Academy of Agricultural Sciences/Jiangsu Technical Service Center of Diagnosis and Detection for Plant Virus Diseases, Nanjing 210014, China;

Received:2013-04-27 Revised:2013-05-17 Online:2013-11-10 Published:2013-11-10
Contact: ZHOU Yijun*

摘要/Abstract

摘要： 为获取灰飞虱体内传播RBSDV相关介体因子，以RBSDV编码的外层外壳蛋白p10为诱饵，采用酵母双杂交方法从灰飞虱cDNA文库中筛选与RBSDV p10互作的蛋白。将RBSDV p10 基因构建至pGBKT7载体获得诱饵表达载体pGBKT7p10。自激活实验结果表明，p10蛋白对酵母细胞无毒性，也不具有自激活活性。通过pGADT7cDNA文库质粒转化含有pGBKT7p10的酵母AH109筛选获得灰飞虱cDNA文库中与p10互作的蛋白。文库筛选共获得326个阳性克隆。测序结果表明这些阳性克隆编码14种不同的互作蛋白，包括Actin 1，GAPDH，RACK等。这些互作蛋白参与胞吞胞吐作用和膜融合等过程，可能与病毒在介体内的循回和增殖相关。

关键词: 水稻黑条矮缩病毒, 外壳蛋白, 互作蛋白, 酵母双杂交

Abstract: To reveal protein factors in Laodelphax striatellus Fallen(the small brown planthopper, SBPH) that related to Rice blackstreaked dwarf virus (RBSDV) transmission, the outer capsid protein p10 encoded by RBSDV S10 was selected as bait to screen the SBPH cDNA library. RBSDV p10 gene was inserted to pGBKT7 vector and the bait plasmid pGBKT7p10 was constructed. The result of autoactivation test showed that p10 could not autonomously activate the expression of reporter genes in yeast and had no toxicity to yeast cell. To obtain the interacting proteins with p10, the pGADT7cDNA library plasmids were transformed to the yeast AH109 which contains pGBKT7p10. Three hundred and twentysix positive clones were acquired. The sequencing results show that these positive clones encode 14 proteins, including Actin 1, GAPDH and RACK. These interacting proteins were involved in several processes such as endocytosis, exocytosis and membrane fusion. The interactions between p10 and these proteins may be related to the viral circulation and proliferation in insect vector.

Key words: Rice blackstreaked dwarf virus, capsid protein, interacting protein, yeast two hybrid

中图分类号:

徐秋芳,陈晴晴,张金凤,李硕,倪海平,周益军* . 酵母双杂交筛选灰飞虱体内水稻黑条矮缩病毒 p10 的互作蛋白[J]. 中国水稻科学, 2013, 27(6): 633-638.

XU Qiufang, CHEN Qingqing, ZHANG Jinfeng, LI Shuo, NI Haiping, ZHOU Yijun*. Screening of RBSDV p10 Interacting Proteins in Small Brown Planthoppers by Yeast Two Hybrid System[J]. Chinese Journal of Rice Science, 2013, 27(6): 633-638.

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酵母双杂交筛选灰飞虱体内水稻黑条矮缩病毒 p10 的互作蛋白

Screening of RBSDV p10 Interacting Proteins in Small Brown Planthoppers by Yeast Two Hybrid System

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