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Cloning and expression of Spodoptera litura nucleopolyhedrovirus gp41 gene in Escherichia coli and preparation of antibody

Published online by Cambridge University Press:  13 June 2008

Pan Li-Jing ,
Li Zhao-Fei ,
Yin Chong ,
Lv Lei  and
Pang Yi
Pan Li-Jing
Affiliation:
State Key Laboratory for Biocontrol, Zhongshan University, Guangzhou 510275, China
Li Zhao-Fei
Affiliation:
State Key Laboratory for Biocontrol, Zhongshan University, Guangzhou 510275, China
Yin Chong
Affiliation:
State Key Laboratory for Biocontrol, Zhongshan University, Guangzhou 510275, China
Lv Lei
Affiliation:
State Key Laboratory for Biocontrol, Zhongshan University, Guangzhou 510275, China
Pang Yi*
Affiliation:
State Key Laboratory for Biocontrol, Zhongshan University, Guangzhou 510275, China
*
*Corresponding author. Email: [email protected]
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Abstract

GP41, a major glycoprotein, identified in the occlusion-derived virions (ODV) of baculoviruses, is required for the egress of nucleocapsids from the nucleus in the pathway of budded virion (BV) synthesis. Using the polymerase chain reaction (PCR), the open reading frame (ORF) of Spodoptera litura nucleopolyhedrovirus (SpltMNPV) gp41 gene was obtained from SpltMNPV genomic DNA. The PCR product was cloned into pMD18-T vector to get the recombinant plasmid (pT-gp41). The gp41 gene was recombined in vitro with prokaryotic expression vector pQE30 and transformed into Escherichia coli M15 [pREP4]. The M15 [pREP4] strain, containing gp41 recombinant plasmid, expressed a 37.9 kDa 6×His-tag fusion protein after induction with 1 mmol/l isopropylthio-β-d-galactoside (IPTG). The fusion protein was purified with a nickel-nitrilotriacetic acid (Ni–NTA) resin column and used as the immunogen to raise GP41-specific antibody. Western blotting analysis indicated that the antibody was suitable to be used for further analysis of GP41 protein.

Keywords

Spodoptera litura nucleopolyhedrovirusgp41antibody
Type
Research Article
Information
Chinese Journal of Agricultural Biotechnology , Volume 2 , Issue 2 , August 2005 , pp. 113 - 117
Copyright
Copyright © China Agricultural University and Cambridge University Press 2005

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References

Herniou, EA, Olszewski, JA, Cory, JS et al. , (2003) The genome sequence and evolution of baculoviruses. Annual Review of Entomology 48: 211–234.CrossRefGoogle ScholarPubMed
Li, GH, Chen, QJ and Pang, Y (1988) Studies on artificial diets for the beet armyworm, Spodoptera exigua. Acta Scientiarum Naturalium Universitatis SunYatSeNi 37 (4): 1–5.Google Scholar
Miller, LK (1997) Introduction to the baculovirus. In: Miller, LK (editor) The Baculovirus. New York: Plenum Press, pp. 1–6.CrossRefGoogle ScholarPubMed
Olszewski, J and Miller, LK (1997) A role for baculovirus GP41 in budded virus production. Virology 33 (2): 292–301.CrossRefGoogle Scholar
O'Reilly, DR, Miller, LK and Luckow, VA (1992) Baculovirus Expression Vectors – A Laboratory Manual. New York: W.H. Freeman and Co., pp. 139–179.Google Scholar
Pang, Y, Yu, JX, Wang, LH et al. , (2001) Sequence analysis of the Spodoptera litura multicapsid nucleopolyhedrovirus genome. Virology 287: 391–404.CrossRefGoogle ScholarPubMed
Sambrook, J, Fritsch, EF and Maniatis, T (1989) Molecular Cloning: A Laboratory Manual. New York: Cold Spring Harbor Laboratory Press.Google Scholar
Volkman, LE (1986) The 64K envelope protein of budded Autographa californica nuclear polyhedrosis virus. Current Topics in Microbiology and Immunology 131: 103–118.Google ScholarPubMed
Whitford, M and Faulkner, P (1992) A structural polypeptide of the baculovirus Autographa californica nuclear polyhedrosis virus contains O-linked N-acetylglucosamine. Journal of Virology 66: 3324–3329.CrossRefGoogle ScholarPubMed
Xie, WD, Qu, SR, Pang, Y et al. , (1988) Establishment of a cell line from Spodoptera litura and its susceptibility to nuclear polyhedrosis viruses. Acta Scientiarum Naturalium Universitatis SunYatSeNi 4: 113–116.Google Scholar