Prokaryotic expression, purification and identification of the N protein of middle east respiratory syndrome coronavirus
10.3760/cma.j.issn.0254-5101.2014.10.007
- VernacularTitle:中东呼吸综合征冠状病毒N蛋白的原核表达、纯化和鉴定
- Author:
Yanqun WANG
;
Chunxu HAO
;
Yao DENG
;
Yang YANG
;
Li ZHAO
;
Xiaoling SHEN
;
Wenjie TAN
- Publication Type:Journal Article
- Keywords:
Middle east respiratory syndrome coronavirus;
N protein;
Prokaryotic expression;
Purification;
Identification
- From:Chinese Journal of Microbiology and Immunology
2014;(10):764-769
- CountryChina
- Language:Chinese
-
Abstract:
Objective To express the N protein of middle east respiratory syndrome coronavirus ( MERS-CoV) in prokaryotic expression system and evaluate the immunogenicity of the purified recombinant N protein.Methods The gene encoding N protein of MERS-CoV was synthesized and cloned into the vector pET30a to construct the recombinant expression plasmid pET30a-MERS-CoV-N.The transformed E.coli BL21 ( DE3) strains carrying expression plasmid were induced by IPTG to express N protein.The expressed protein was purified by using affinity chromatography.SDS-PAGE and Western blot assays were used to iden-tify the expressed N protein and evaluate its immunogenicity.Results The recombinant N protein was suc-cessfully expressed in soluble form with the size of 46×103 .The results of Western blot assay showed that the recombinant N protein could specifically react with rabbit serum samples positive for antibodies against N protein B-cell epitope peptide and mouse anti-His tag antibodies.No positive band against N protein was found when primary antibody was used with thirty adult serum samples from Beijing in 2008.Conclusion N protein of MERS-CoV was successfully expressed in prokaryotic expression system.The successful expres-sion of N protein laid the foundation for immunological diagnosis of N protein of MERS-CoV and researches on its immunogenicity.
