[1]郭运泽?,孙恩成?,徐青元,等.蓝舌病病毒10质粒反向遗传操作系统的建立[J].中国预防兽医学报,2019,(03):234-238.[doi:10.3969/j.issn.1008-0589.201807005]
 GUO Yun-ze,SUN En-cheng,XU Qing-yuan,et al.Establishment of a ten plasmid-based reverse genetics system for Bluetongue virus[J].Chinese journal of preventive veterinary medicine,2019,(03):234-238.[doi:10.3969/j.issn.1008-0589.201807005]
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蓝舌病病毒10质粒反向遗传操作系统的建立()
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《中国预防兽医学报》[ISSN:1008-0589/CN:23-1417/S]

卷:
期数:
2019年03
页码:
234-238
栏目:
病原生物学
出版日期:
2019-04-25

文章信息/Info

Title:
Establishment of a ten plasmid-based reverse genetics system for Bluetongue virus
文章编号:
1008-0589(2019)03-0234-05
作者:
 

郭运泽12?孙恩成2?徐青元2步志高2吴东来2*王凤龙12*

 (1. 内蒙古农业大学 兽医学院,内蒙古 呼和浩特 010018;2. 中国农业科学院哈尔滨兽医研究所 兽医生物技术国家重点实验室/
农业部兽医公共卫生重点开放实验室,黑龙江 哈尔滨 150069)
Author(s):
 

GUO Yun-ze12? SUN En-cheng2? XU Qing-yuan2  BU Zhi-gao2 WU Dong-lai2* WANG Feng-long12*

 (1. College of Veterinary Medicine, Inner Mongolia Agricultural University, Hohhot 010018, China; 2. State Key Laboratory of Veterinary Biotechnology, Veterinary Public Health Laboratory of Ministry of Agricultural Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150069, China)
关键词:
蓝舌病病毒反向遗传质粒系统
Keywords:
Bluetongue virus  reverse genetics  plasmid-based system
分类号:
S852.65
DOI:
10.3969/j.issn.1008-0589.201807005
文献标志码:
A
摘要:
为建立蓝舌病病毒(BTV) 10质粒反向遗传操作系统,本研究将BTV16型BN96/16株10个基因节段分别克隆至pS111载体相应的酶切位点,构建10个重组质粒转染BSR细胞,拯救获得重组病毒,命名为rBTV-16。测序结果显示,拯救病毒rBTV-16与其亲本病毒wtBTV-16的核苷酸序列完全相同;经电镜观察,可见典型形态的环状病毒属病毒粒子;间接免疫荧光试验结果显示,rBTV-16能够感染细胞;病毒基因组检测结果显示,rBTV-16各个基因节段的长度与亲本病毒完全一致;病毒生长曲线的测定结果显示,rBTV-16与亲本病毒具有一致的复制特性。本研究建立的10质粒系统比传统体外转录拯救系统的操作过程简便,提高了拯救病毒的效率,为进一步深入研究BTV的致病机理奠定了基础。
  
Abstract:
To establish an entirely plasmid-based reverse genetics system for Bluetongue virus (BTV), ten reverse genetics plasmids were constructed by 10 segments of BTV-16 BN96/16 strain genome cloning into vector pS111. The recombinant BTV-16 (rBTV-16) was successfully rescued from BSR cells after transfecting with the 10 plasmids. The nucleotide sequences of rBTV-16 were identical to wtBTV-16, which was confirmed by sequencing the respective genome segments and the typical Orbivirus virions were observed by electron microscopy. The cells were infected with rBTV-16 confirmed by indirect immunofluorescent assay. The genome dsRNA was extracted and analyzed by non-denaturing polyacrylamide gel electrophoresis. The growth curves indicated that rBTV-16 had similar replication characteristics to its parental virus of wtBTV-16. The 10 plasmid-based system greatly simplifies the complex progress of in vitro transcription system, and also improves the efficiency of virus rescue, which provides a promising technique support for further study on the pathogenic of BTV.

参考文献/References:

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                                (本文编辑:彭永刚)

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更新日期/Last Update: 2019-04-26