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题名: Rana grylio virus as a vector for foreign gene expression in fish cells
作者: He, Li-Bo1; Ke, Fei1; Zhang, Qi-Ya1
通讯作者: Zhang, QY (reprint author), Chinese Acad Sci, Grad Sch, Inst Hydrobiol, State Key Lab Freshwater Ecol & Biotechnol, Wuhan 430072, Peoples R China.
关键词: Viral vector ; Recombinant iridovirus ; Rana grylio virus (RGV) ; Thymidine kinase (TK) ; Viral envelope protein 53R ; Epithelioma papulosum cyprinid (EPC)
刊名: VIRUS RESEARCH
发表日期: 2012
DOI: 10.1016/j.virusres.2011.08.012
卷: 163, 期:1, 页:66-73
收录类别: SCI
文章类型: Article
部门归属: [He, Li-Bo; Ke, Fei; Zhang, Qi-Ya] Chinese Acad Sci, Grad Sch, Inst Hydrobiol, State Key Lab Freshwater Ecol & Biotechnol, Wuhan 430072, Peoples R China
WOS标题词: Science & Technology ; Life Sciences & Biomedicine
资助者: National Major Basic Research Program[2009CB118704, 2010CB126303]; National Natural Science Foundation of China[30871938, 31072239]; Chinese Academy of Sciences[KSCX2-EW-Z-3]; State Key Laboratory of Freshwater Ecology and Biotechnology[2008FBZ15, 2008FBZ16]
类目[WOS]: Virology
研究领域[WOS]: Virology
摘要: In the present study, Rana grylio virus (RGV, an iridovirus) thymidine kinase (TK) gene and viral envelope protein 53R gene were chosen as targets for foreign gene insertion. Delta TK-RGV and Delta 53R-RGV, two recombinant RGV, expressing enhanced green fluorescence protein (EGFP) were constructed and analyzed in Epithelioma papulosum cyprinid (EPC) cells. The EGFP gene which fused to the virus major capsid protein (MCP) promoter p50 was inserted into TK and 53R gene loci of RGV, respectively. Cells infected with these two recombinant viruses not only displayed plaques, but also emitted strong green fluorescence under fluorescence microscope, providing a simple method for selection and purification of recombinant viruses. Delta TK-RGV was purified by seven successive rounds of plaque isolation and could be stably propagated in EPC cells. All of the plaques produced by the purified recombinant virus emitted green fluorescence. However, Delta 53R-RGV was hard to be purified even through twenty rounds of plaque isolation. The purified recombinant virus Delta TK-RGV was verified by PCR analysis and Western blotting. These results showed EGFP was expressed in Delta TK-RGV infected cells. Furthermore, one-step growth curves and electron microscopy revealed that infection with recombinant Delta TK-RGV and wild-type RGV are similar. Therefore, RGV was demonstrated could be as a viral vector for foreign gene expression in fish cells. Crown Copyright (C) 2011 Published by Elsevier B.V. All rights reserved.
英文摘要: In the present study, Rana grylio virus (RGV, an iridovirus) thymidine kinase (TK) gene and viral envelope protein 53R gene were chosen as targets for foreign gene insertion. Delta TK-RGV and Delta 53R-RGV, two recombinant RGV, expressing enhanced green fluorescence protein (EGFP) were constructed and analyzed in Epithelioma papulosum cyprinid (EPC) cells. The EGFP gene which fused to the virus major capsid protein (MCP) promoter p50 was inserted into TK and 53R gene loci of RGV, respectively. Cells infected with these two recombinant viruses not only displayed plaques, but also emitted strong green fluorescence under fluorescence microscope, providing a simple method for selection and purification of recombinant viruses. Delta TK-RGV was purified by seven successive rounds of plaque isolation and could be stably propagated in EPC cells. All of the plaques produced by the purified recombinant virus emitted green fluorescence. However, Delta 53R-RGV was hard to be purified even through twenty rounds of plaque isolation. The purified recombinant virus Delta TK-RGV was verified by PCR analysis and Western blotting. These results showed EGFP was expressed in Delta TK-RGV infected cells. Furthermore, one-step growth curves and electron microscopy revealed that infection with recombinant Delta TK-RGV and wild-type RGV are similar. Therefore, RGV was demonstrated could be as a viral vector for foreign gene expression in fish cells. Crown Copyright (C) 2011 Published by Elsevier B.V. All rights reserved.
关键词[WOS]: SINGAPORE GROUPER IRIDOVIRUS ; SWINE-FEVER VIRUS ; CHILO-IRIDESCENT-VIRUS ; MAJOR CAPSID PROTEIN ; COMPLETE GENOME SEQUENCE ; FAMILY-IRIDOVIRIDAE ; HUMAN CYTOMEGALOVIRUS ; PROTEOMIC ANALYSIS ; MEMBRANE-PROTEIN ; BOHLE IRIDOVIRUS
语种: 英语
WOS记录号: WOS:000301160200008
ISSN号: 0168-1702
Citation statistics:
内容类型: 期刊论文
URI标识: http://ir.ihb.ac.cn/handle/342005/16957
Appears in Collections:鱼类生物学及渔业生物技术研究中心_期刊论文

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作者单位: 1.Chinese Acad Sci, Grad Sch, Inst Hydrobiol, State Key Lab Freshwater Ecol & Biotechnol, Wuhan 430072, Peoples R China

Recommended Citation:
He, Li-Bo; Ke, Fei; Zhang, Qi-Ya.Rana grylio virus as a vector for foreign gene expression in fish cells,VIRUS RESEARCH,2012,163(1):66-73
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