IHB OpenIR  > 鱼类生物学及渔业生物技术研究中心  > 期刊论文
Rana grylio virus as a vector for foreign gene expression in fish cells
He, Li-Bo; Ke, Fei; Zhang, Qi-Ya; Zhang, QY (reprint author), Chinese Acad Sci, Grad Sch, Inst Hydrobiol, State Key Lab Freshwater Ecol & Biotechnol, Wuhan 430072, Peoples R China.
2012
Source PublicationVIRUS RESEARCH
ISSN0168-1702
Volume163Issue:1Pages:66-73
AbstractIn 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.; 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.
SubtypeArticle
KeywordViral Vector Recombinant Iridovirus Rana Grylio Virus (Rgv) Thymidine Kinase (Tk) Viral Envelope Protein 53r Epithelioma Papulosum Cyprinid (Epc)
Department[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
DOI10.1016/j.virusres.2011.08.012
WOS HeadingsScience & Technology ; Life Sciences & Biomedicine
Funding OrganizationNational 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] ; 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] ; 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] ; 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]
Indexed BySCI
Language英语
WOS Research AreaVirology
WOS SubjectVirology
WOS IDWOS:000301160200008
WOS KeywordSINGAPORE GROUPER IRIDOVIRUS ; SWINE-FEVER VIRUS ; CHILO-IRIDESCENT-VIRUS ; MAJOR CAPSID PROTEIN ; COMPLETE GENOME SEQUENCE ; FAMILY-IRIDOVIRIDAE ; HUMAN CYTOMEGALOVIRUS ; PROTEOMIC ANALYSIS ; MEMBRANE-PROTEIN ; BOHLE IRIDOVIRUS
Funding OrganizationNational 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] ; 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] ; 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] ; 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]
Citation statistics
Cited Times:11[WOS]   [WOS Record]     [Related Records in WOS]
Document Type期刊论文
Identifierhttp://ir.ihb.ac.cn/handle/342005/16957
Collection鱼类生物学及渔业生物技术研究中心_期刊论文
Corresponding AuthorZhang, QY (reprint author), Chinese Acad Sci, Grad Sch, Inst Hydrobiol, State Key Lab Freshwater Ecol & Biotechnol, Wuhan 430072, Peoples R China.
AffiliationChinese Acad Sci, Grad Sch, Inst Hydrobiol, State Key Lab Freshwater Ecol & Biotechnol, Wuhan 430072, Peoples R China
Recommended Citation
GB/T 7714
He, Li-Bo,Ke, Fei,Zhang, Qi-Ya,et al. Rana grylio virus as a vector for foreign gene expression in fish cells[J]. VIRUS RESEARCH,2012,163(1):66-73.
APA He, Li-Bo,Ke, Fei,Zhang, Qi-Ya,&Zhang, QY .(2012).Rana grylio virus as a vector for foreign gene expression in fish cells.VIRUS RESEARCH,163(1),66-73.
MLA He, Li-Bo,et al."Rana grylio virus as a vector for foreign gene expression in fish cells".VIRUS RESEARCH 163.1(2012):66-73.
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