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水稻黑条矮缩病毒外壳蛋白基因的克隆、原核表达及抗体制备

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Keywords: 水稻黑条矮缩病毒,外壳蛋白,序列分析,原核表达,多克隆抗体

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Abstract:

为探讨水稻黑条矮缩病毒(Riceblack-streakeddwarfvirus,RBSDV)的种群变异,采用RT-PCR方法从感染RBSDV山东分离物RBSDV-JN1的水稻中克隆该病毒的S10片段,并进行序列分析,进而构建包含RBSDV-JN1的CP基因的原核表达载体,导入大肠杆菌EscherichiacoliBL21(DE3)诱导表达;并以表达的融合蛋白为抗原,制备病毒的多克隆抗体。结果显示:S10片段全长为1801bp,包含1个1677bp编码框,编码558个氨基酸的外壳蛋白(CP);与GenBank已注册的19个RBSDV的CP基因序列相比较发现,病毒各分离物间核苷酸的序列相似性为90.5%~99.8%,氨基酸的序列相似性为95.9%~100.0%;地理位置较近的分离物间序列相似性较高,RBSDV种群分布呈现区域性差异。原核表达载体pET-RBSDV-CP经IPTG诱导,获得了分子量约为63kD带有His标签的目的蛋白。用该蛋白制备的多克隆抗体经ELISA、Westernblot和Dot-blotELISA检测显示,效价为1:81000,且具有良好的特异性。

References

[1]  Bai F W, Yan J, Qu Z C, et al. Phylogenetic analysis reveals that a dwarfing disease on different cereal crops in China is due to rice black streaked dwarf virus (RBSDV). Virus Genes, 2002, 25(2): 201-206
[2]  Chen Shengxiang, Zhang Qiaoyan. Advance in researches on rice black-streaked dwarf disease and maize rough dwarf disease in China. Journal of Plant Protection, 2005, 32(1): 98-103 (in Chinese) [陈声祥, 张巧艳. 我国水稻黑条矮缩病和玉米粗缩病研究进展. 植物保护学报, 2005, 32(1): 98-103]
[3]  Ling K C. Rice virus diseases. Philippines: International Rice Research Institute, 1972
[4]  Long Mengling. Rice black streaked dwarf disease and its prevention and treatment research progress in our country. Guangxi Plant Protection, 2011, 24(1): 30-32 (in Chinese) [龙梦玲. 我国水稻黑条矮缩病及其防治研究进展. 广西植保, 2011, 24(1): 30-32]
[5]  Azuhata F, Uyeda I, Kimura I, et al. Close similarity between genome structures of rice black streaked dwarf and maize rough dwarf viruses. Journal of General Virology, 1993, 74(7): 1227-1232
[6]  Shikata E, Kitagawa Y. Rice black-streaked dwarf virus: its properties, morphology and intracellular localization. Virology, 1977, 77(2): 826-842
[7]  Zhang H M, Chen J P, Adams M J. Molecular characterization of segments 1 to 6 of Rice black streaked dwarf virus from China provides the complete genome. Archives of Virology, 2001, 146(12): 2331-2339
[8]  Wang Z H, Fang S G, Xu J L, et al. Sequence analysis of the complete genome of Rice black streaked dwarf virus isolated from maize with rough dwarf disease. Virus Genes, 2003, 27(2): 163-168
[9]  Zhang Hengmu, Chen Jianping, Xue Qingzhong. cDNA cloning and sequence analysis of genome segment s10 from rice black-streaked dwarf virus. Chinese Journal of Rice Science, 2002, 16(1): 24-28 (in Chinese) [张恒木, 陈剑平, 薛庆中, 等. 水稻黑条矮缩病毒基因组片段S10的cDNA克隆及全序列分析. 中国水稻科学, 2002, 16(1): 24-28]
[10]  Wang Zhaohui, Zhou Yijun, Fan Yongjian, et al. cDNA clone and sequence analysis of segment 10 of rice black-streaked dwarf fijivirus in jiangsu. Virologica Sinica, 2002, 17(2): 142-144 (in Chinese) [王朝辉, 周益军, 范永坚, 等. 江苏水稻黑条矮缩病毒S10的cDNA克隆序列分析. 中国病毒学, 2002, 17(2): 142-144]
[11]  Bai Fengwei, Qu Zhicai, Cao Qingyu, et al. Cloning and expression of rice black streaked dwarf virus genome components 10 coat protein coding genes. Journal of Fudan University, 2002, 41(2): 217-221 (in Chinese) [白逢伟, 曲志才, 曹清玉, 等. 水稻黑条矮缩病毒基因组组分10编码的外壳蛋白基因的克隆及表达. 复旦学报, 2002, 41(2): 217-221]
[12]  Fang S, Yu J, Feng J, et al. Identification of rice black streaked dwarf fijivirus in maize with rough dwarf disease in China. Archives of Virology, 2001, 146(1): 167-170
[13]  Isogai M, Uyeda I, Lee B C. Detection and assignment of proteins encoded by rice black streaked dwarf fijivirus S7, S8, S9 and S10. Journal of General Virology, 1998, 79(6): 1487-1494
[14]  Deng Jinqi, Dai Liangying. Research progress on rice black streaked dwarf virus (RBSDV). Chinese Agricultural Science Bulletin, 2012, 28(18): 209-212 (in Chinese) [邓金奇, 戴良英. 水稻黑条矮缩病毒的研究进展. 中国农学通报, 2012, 28(18): 209-212]
[15]  Yang Jian, Zhang Hengmu, Chen Jianping, et al. Prekaryotic expression antiserum preparation and some properties of p8 protein of rice black-streaked dwarf Fijivirus. Journal of Plant Protection, 2007, 34(3): 252-256 (in Chinese) [羊健, 张恒木, 陈剑平, 等. 水稻黑条矮缩病毒p8蛋白的原核表达、抗血清制备及其特性. 植物保护学报, 2007, 34(3): 252-256]
[16]  Ouyang Yuanlong Wu Jianxiang Xiong Ruyi, et al. Prokaryotic expression of coat protein gene S10 of rice black-streaked dwarf virus,and preparation and application of its polyclonal antibody. Chinese Journal of Rice Science, 2010, 24(1): 25-30 (in Chinese) [欧阳元龙, 吴建祥, 熊如意, 等. 水稻黑条矮缩病病毒外壳蛋白基因S10的原核表达、多克隆抗体制备及应用. 中国水稻科学, 2010, 24(1): 25-30]
[17]  Sambrook J, fees Ridge EF, Maniatis T (translated by Huang Peitang, et al). Laboratory guidelines of molecular cloning(3 edition). Beijing: Science Press, 2002 (in Chinese) [萨姆布鲁克 J, 费里奇 E F, 曼尼阿蒂斯 T. (黄培堂等译). 分子克隆实验指南(3版). 北京: 科学出版社, 2002]
[18]  Wu J X, Yu L, Li L, et al. Oral immunization with transgenic rice seeds expressing VP2 protein of infectious bursal disease virus induces protective immune responses in chickens. Plant Biotechnology Journal, 2007, 5(5): 570-578
[19]  Zhang Haiyan, Diao Yonggang, Yang Haibo , et al. Population dynamics and migration characteristics of the small brown planthopper in spring in Jining,Shandong Province. Chinese Journal of Applied Entomology, 2011, 48(5): 1298-1308 (in Chinese) [张海燕, 刁永刚, 杨海博, 等. 山东济宁灰飞虱春季种群动态及迁飞特性. 应用昆虫学报, 2011, 48(5): 1298-1308]
[20]  Ren Zengliang, Du Guocheng, Chen Jian , et al. Strategies for high-level expression of recombinant protein in Escherichia coli. China Biotechnology, 2007, 27(9): 103-109 (in Chinese) [任增亮, 堵国成, 陈坚, 等. 大肠杆菌高效表达重组蛋白策略. 中国生物工程杂志, 2007, 27(9): 103-109]
[21]  更多...
[22]  Chen Mingjie, Gao Mingming, Yao Wenbing, et al. Influence on the expression of heterologous proteins in Escherichia coliofnonop timal codon quantities and location. Journal of China Pharmaceutical University, 2008, 39(6): 553-556 (in Chinese) [陈明杰, 高明明, 姚文兵, 等. 大肠杆菌非偏爱密码子的数量及位置对外源蛋白表达量的影响. 中国医药大学学报, 2008, 39(6): 553-556]
[23]  Gustafsson C, Govindarajan S, Minshull J. Codon bias and heterologous protein expression. Trends in Biotechnology, 2004, 22(7): 346-353
[24]  de Valdivia E I G, Isaksson L A. A codon window in mRNA downstream of the initiation codon where NGG codons give strongly reduced gene expression in Escherichia coli. Nucleic Acids Research, 2004, 32(17): 5198-5205

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