鸡传染性支气管炎病毒(IBV)ZZ2004株核衣壳蛋白基因的克隆及原核表达

Abstract

Abstract: This experiment was conducted to clone and express the nucleoprotein (N) gene of infectious bronchitis virus (IBV). A pair of specific primers was designed and synthesized according to the published sequence of IBV ZZ2004. RNA of IBV ZZ2004 strain was extracted and the fragment of about 1.23 kb was amplified by RT-PCR, which was correct fragment.The fragment was inserted into the cloning vector pGEM-T to construct pGEM-T-N, and nucleotide sequence was determined by dideoxy-mediated chain termination method. The pGEM-T-N was excised by EcoR Ⅰ and Xho Ⅰ, and inserted into the expression plasmid pGEX-6P-1 to obtain the recombinant expression vector pGEX-6P-1-N, which was used to transform into E.coli BL21 (DE3) competent cells. Then the recombinant pGEX-6P-1-N was induced by IPTG, SDS-PAGE and Western blotting results confirmed that the gene had been expressed in recombinant pGEX-6P-1-N, and the protein was soluble. The expression product could specially react to the anti-sera of IBV which showed that N protein had some biological activity. This study had laid a solid foundation for production of diagnostic reagents for testing IB and new type IB gene engineering vaccines.

Key words: infectious bronchitis virus(IBV); nucleoprotein; cloning; soluble expression

CLC Number:

S852.65

HE Hui-li, XU Min, YU Juan, WEI Xiao-bing, YAN Yi-ting, GOU Xiao-jing, LIU Xing-you. Cloning and Prokaryotic Expression of Nucleoprotein Gene of Infectious Bronchitis Virus (IBV)ZZ2004 Strain[J]. , 2014, 41(9): 15-20.

References

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Cloning and Prokaryotic Expression of Nucleoprotein Gene of Infectious Bronchitis Virus (IBV)ZZ2004 Strain

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