Prokaryotic Expression and Purification of Nonstructural Protein NS1 of Goose Tembusu Virus

doi:10.3969/j.issn.1000-7091.2012.06.003

Abstract

Abstract: The goose tembusu virus JS804 strain non-structural protein NS1 gene was amplified by PCR with the specific primers designed on the basis of NS1 gene and then was inserted into the prokaryotic vector pET28a and pET32a for the construction of recombinant expression plasmid pET28a-NS1 and pET32a-NS1． Then pET28a-NS1 and pET32a-NS1 were transformed respectively into Escherichia coli BL21 ( DE3). The recombinant protein NS1s ( His-NS1) were obtained with the induction of IPTG and the molecular weight of the fusion protein was 44 kDa and 58 kDa respectively． After 6 h induction by IPTG， the yield of fusion proteins reached peaks． The analysis showed that two fusion proteins were expressed in Escherichia coli into inclusion bodies． The fusion protein was purified to pure and high-level proteins by extracting the inclusion bodies with urea． The results laid the foundation for the further studies on the NS1 protein of goose tembusu virus．

Key words: Goose tembusu virus, NS1 protein, Prokaryotic expression

CLC Number:

ZHAO Dong-min, HUANG Xin-mei, LIU Yu-zhuo, ZHANG Jing-feng, HAN Kai-kai, XIE Xing-xing, ZHOU Xiao-bo, LI Yin. Prokaryotic Expression and Purification of Nonstructural Protein NS1 of Goose Tembusu Virus[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2012, 27(6): 11-14. doi: 10.3969/j.issn.1000-7091.2012.06.003.

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References

[1] 黄欣梅，李银，赵冬敏，等．新型鹅黄病毒JS804 毒株的分离与鉴定[J]．江苏农业学报， 2011， 27( 2) : 354-360．
[2] 苏敬良．鸭的新型黄病毒BYD 引起的产蛋下降综合征[J]．兽医导刊， 2011( 4) : 27-29．
[3] Cao Z，Zhang C，Liu Y， et al． Tembusu virus in ducks，China[J]． Emerg Infect Dis， 2011， 17( 10) : 1873-1875．
[4] 张大丙．鸭出血性卵巢炎的研究进展[J]．中国家禽， 2011， 33( 14) : 37-38．
[5] Yun T，Ye W，Ni Z，et al． Identification and molecular characterization of a novel flavivirus isolated from Pekin ducklings in China[J]． Vet Microbiol， 2012， 157( 3 /4) : 311-319．
[6] 姬希文，李雪松，李国新，等．鸭坦布苏病毒E 蛋白中和性单克隆抗体的制备[J]．中国家禽，2012，34( 4) :25-32．
[7] MuylaeRTI R，Galler R，Rice C M． Genetic analysis of the yellow fever virus NS1 protein: identification of a temperature-sensitive mutation which blocks RNA accumulation[J]． J Virol， 1997， 71( 1) : 291-298．
[8] Lindenbach B D，Rice C M． Genetic interaction of flavivirus nonstructural proteins NS1 and NS4A as a determinant of replicase function[J]． J Virol， 1999， 73: 4611-4621．
[9] Lindenbach B D，Rice C M． Trans-complementation of yellow fever virus NS1 reveals a role in early RNA replication[J]． J Virol， 1997， 71: 9608-9617．
[10] Pacca C C，Severino A A，Mondini A， et al． RNA interference inhibits yellow fever virus replication in vitro and vivo[J]． Virus genes， 2009， 38( 2) 224-231．
[11] Alcon-LePoder S， Sivard P，Drouet M T， et al． Secretion of flaviviral non-structural protein NS1: from diagnosis to pathogenesis[J]． Novartis Found Symp， 2006， 277:233-247．
[12] Avirutnan P，Punyadee N，Noisakran S， et al． Vascular leakage in severe dengue virus infections: a potential role for the nonstructural viral protein NS1 and complement[J]． J Infect Dis， 2006， 193( 8) : 1078-1088．
[13] Schlesinger J J． Flavivirus nonstructural protein NS1: complementary surprises[J]． Proc Natl Acad Sci USA， 2006， 103( 50) : 18879-18880．
[14] Tafuku S，Miyata T，Tadano M， et al． Japanese encephalitis virus structural and nonstructural proteins expressed in Escherichia coli induce protective immunity in mice[J]． Microbes Infect， 2012， 14( 2) : 169-176．
[15] Mackenzie J M， Jones M K，Young P R． Immunolocalization of the dengue virus nonstructural glycoprotein NS1 suggests a role in viral RNA replication[J]． Virology， 1996， 220: 232-240．
[16] Konishi E，Kitai Y． Detection by ELISA of antibodies to Japanese encephalitis virus nonstructural 1 protein induced in subclinically infected humans[J]． Vaccine， 2009， 27( 50) : 7053-7058．

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