SRBSDV侵染的白背飞虱中肠酵母双杂交cDNA文库的构建和分析

摘要
图/表
参考文献
相关文章 (2)

全文: PDF (2993 KB) HTML (1 KB)
输出: BibTeX | EndNote (RIS)

摘要南方水稻黑条矮缩病毒(Southern rice black-streaked dwarf virus, SRBSDV)是呼肠孤病毒科(Reoviridae)斐济病毒属(Fijivirus)成员，主要由白背飞虱(Sogatella furcifera, WBPH)以持久增殖型方式进行传播。在病毒的侵染循回过程中，白背飞虱中肠上皮细胞是病毒的初侵染和主要增殖场所。而病毒的有效增殖是决定介体能否传毒的关键影响因素。为了更好地研究SRBSDV和介体白背飞虱的互作关系，尤其是了解白背飞虱中肠蛋白通过参与调控病毒的增殖过程，而使介体昆虫成功获毒并传毒，本研究以高带毒白背飞虱群体中肠组织为实验材料，构建了高带毒白背飞虱群体中肠的酵母双杂交cDNA文库。经过检测表明，构建的文库滴度为1.5×106 cfu/mL，平均插入片段主要分布在1.0~2.0 kb之间，文库质量较好，可用于研究SRBSDV编码蛋白和白背飞虱中肠蛋白的互作关系，并为开展SRBSDV和昆虫介体的互作研究奠定了基础。

	服务

	把本文推荐给朋友
	加入我的书架
	加入引用管理器
	E-mail Alert
	RSS
	作者相关文章
	郑立敏
	彭静
	陈建斌
	张松柏
	程菊娥
	张德咏
	刘勇

关键词 ：南方水稻黑条矮缩病毒(SRBSDV), 白背飞虱, 中肠, 酵母双杂cDNA文库

Abstract：Southern rice black-streaked dwarf virus (SRBSDV), the genus Fijivirus in the family Reoviridae, is transmitted mainly by white backed planthopper (Sogatella furcifera, WBPH) in a persistent-propagative manner. The epithelial cells of the midgut is the initially infected and replicated tissue for SRBSDV during the whole infection route in its insect vector. The effective multiplication of the virus is the key factor to determine whether the virus can be transmitted. In this study, to investigate the interaction between SRBSDV and its insect vector, and the mechanism of midgut protein regulating viral proliferation to transmit virus successfully. The yeast two-hybrid cDNA library for the midgut of SRBSDV-infected WBPH was constructed through SMART technique. The midgut of WBPH was dissected firstly. The mRNA was isolated and purified from the midguts of viruliferous populations, and used as the template to synthesize the double-strand cDNAs. The cDNAs digested with SfiⅠ were ligated into the library vector pGADT7, and then were transformed into the Escherichia coli DH10B cells to amplify the cDNA library. The cDNA library was calculated the number of transformants per fraction from 1 μL of ligation reaction and the average insert size from the population of 32 released inserts. The results showed the titer of the amplified library was 1.5×106 cfu /mL. The average size of inserts was 1.0~2.0 kb in the cDNA library. The yeast two-hybrid cDNA library derived from the viruliferous WBPH midgut will be useful for the future study on the interaction between insect vector and SRBSDV.

Key words： Southern rice black-streaked dwarf virus (SRBSDV) White backed planthopper Midgut Yeast two-hybrid cDNA library

收稿日期: 2015-08-04 出版日期: 2015-12-29

通讯作者: 张德咏 E-mail: dyzhang78@gmail.com

引用本文:

郑立敏彭静陈建斌张松柏程菊娥张德咏刘勇. SRBSDV侵染的白背飞虱中肠酵母双杂交cDNA文库的构建和分析[J]. , 2016, 24(2): 245-251.

链接本文:

http://journal05.magtech.org.cn/Jwk_ny/CN/ 或 http://journal05.magtech.org.cn/Jwk_ny/CN/Y2016/V24/I2/245

曹杨, 潘峰, 周倩, 等. 2011. 南方水稻黑条矮缩病毒介体昆虫白背飞虱的传毒特性. 应用昆虫学报. 48(5): 1314–1320. (Cao Y, Pan F, Zhou Q, et al. 2011. Transmission characteristics of Sogatella furcifera: A vector of the Southern rice black-streaked dwarf virus. Chinese Journal of Applied Entomology. 48(5): 1314–1320.)
李硕, 孙丽娟, 李醒, 等. 2011. 灰飞虱高带毒( RSV) 群体酵母双杂交cDNA 文库的构建. 昆虫学报. 54(11): 1324–1328. (Li S, Sun L J, Li X, et al. 2011. Construction of yeast two-hybrid cDNA library of high-viruliferous ( RSV-infected) populations of the small brown planthopper, Laodelphax striatellus ( Hemiptera: Delphacidae). Acta Entomologica Sinica. 54(11): 1324–1328.)
沈慧梅. 2010. 我国褐飞虱和白背飞虱的境外虫源研究[D]. 南京: 南京农业大学. (Shen H M. 2010. The study on oversea source area of Nilaparvata lugens and Sogatella furcifera(D). Nanjing: Nanjing Agricultural University.)
王毅. 2007. 白背飞虱的生物学特性. 农技服务. 1: 45–46.
乌慧玲, 许晓凤, 王文兵. 2005. 褐飞虱生物型 I 和生物型 II cDNA文库的构建. 安徽农业大学学报. 32(4): 423–427. (Wu H L, Xu X F, Wang W B. 2005. Full-length cDNA library construction of a specific Tobacco Germplasm. Journal of Anhui Agricultural University. 32(4): 423–427.)
肖冬来, 邓慧颖, 谢荔岩, 等. 2011. 灰飞虱酵母双杂交cDNA文库的构建及分析. 植物保护. 37(1): 19–423. (Xiao D L, Deng H Y, Xie L Y, et al. 2011. Construction of a yeast two-hybrid cDNA library of Laodelphax striatellus. Plant Protection. 37(1): 19–423.)
周国辉, 温锦君, 蔡德江, 等. 2008. 呼肠孤病毒科斐济病毒属一新种: 南方水稻黑条矮缩病毒. 科学通报. 53(20): 2500–2508.
翟保平, 周国辉, 陶小荣, 等. 2011. 稻飞虱暴发与南方水稻黑条矮缩病流行的宏观规律和微观机制. 应用昆虫学报. 48(3): 480–487.(Zhai B P, Zhou G H, Tao X R, et al. 2011. Macroscopic patterns and microscopic mechanisms of the outbreak of rice planthopper and epidemic SRBSDV. Chinese Journal of Applied Entomology. 48(3): 480–487.)
Mar T, Liu W, Wang X. 2014. Proteomic analysis of interaction between P7-1 of Southern rice black-streaked?dwarf?virus?and the insect vector reveals diverse insect proteins involved in successful transmission. Journal of Proteomics. 102: 83–97.
Mao Q Z, Zheng S L, Han Q M, et al. 2013. A new model for the genesis and maturation of the viroplasms induced by plant reoviruses. Journal of Virology. 87(12), 6819–6828.
Haruhiko F, Hajime I. 1986. Molecular mechaniism of introduction of the hidden break into the 28S rRNA of insects: implication based on structural studies. Nucleic Acids Research. 14(16): 6393–6401.
Hogenhout SA, Ammar el-D, Whitfield AE, et al. 2008. Insect vector interactions with persistently transmitted virus. Annual Review of Phytopathology. 46: 327–359.
Jia D S, Chen H Y, Mao Q Z, et al. 2012b. Restriction of viral dissemination from the midgut determines incompetence of small brown planthopper as a vector of Southern rice black-streaked dwarf virus. Virus Research. 167(2): 404–408.
Jia D S, Chen H Y, Zheng A L, et al. 2012a. Development of an insect vector cell culture and RNA interference system to investigate the functional role of fijivirus replication protein. Journal of Virology. 88(10): 5800–5807.
Jia D, Mao Q, Chen H, et al. 2014. Virus-induced tubule: a vehicle for rapid spread of virions through basal lamina from midgut epithelium in the insect vector. Journal of Virology. 88: 10488–10500.
Li J, Xue J, Zhang H, et al. 2015. Characterization of homologous and heterologous interactions between viroplasm proteins P6 and P9-1 of the fijivirus southern rice black-streaked dwarf virus. Archives of Virology. 160(2): 453–457.
Liu Y, Jia D S, Chen H Y, et al. 2011. The P7-1 protein of southern rice black-streaked dwarf virus, a fijivirus, induces the formation of tubular structures in insect cells. Archives of Virology. 156(10): 1729–1736.