Expression and purification of SARS coronavirus membrane protein.
- Author:
Wuxing, DAI
;
Mingjun, LEI
;
Shaoting, WU
;
Zhihao, CHEN
;
Liang, LIANG
;
Huirong, PAN
;
Li, QIN
;
Shitong, GAO
;
Shishan, YUAN
;
Renli, ZHANG
- Publication Type:Journal Article
- MeSH:
Cloning, Molecular;
Escherichia coli/genetics;
Escherichia coli/metabolism;
Membrane Proteins/*biosynthesis;
Membrane Proteins/genetics;
Membrane Proteins/isolation & purification;
Plasmids/biosynthesis;
Plasmids/genetics;
Recombinant Proteins/biosynthesis;
Recombinant Proteins/genetics;
Recombinant Proteins/isolation & purification;
Reverse Transcriptase Polymerase Chain Reaction;
SARS Virus/chemistry;
SARS Virus/*genetics;
Viral Vaccines/biosynthesis
- From:
Journal of Huazhong University of Science and Technology (Medical Sciences)
2004;24(5):414-6
- CountryChina
- Language:English
-
Abstract:
To construct a recombinant plasmid Pet23a-M, the gene encoding severe acute respiratory syndrome (SARS) coronavirus membrane protein was amplified by RT-PCR and cloned into the expression plasmid Pet23a. Results of restriction endonuclease analysis, PCR detection and DNA sequencing analysis revealed that the cloned DNA sequence was the same as that reported. The recombinants were transformed into Escherichia coli (E. Coli) BL21 (DE3) and induced by Isopropyl-beta-D-thiogalactopyranoside (IPTG). The expression of 27 kD (1 kD=0.992 1 ku) protein was detected by SDS-PAGE and pured by metal chelated chromatography. Results of Western-blot showed that this expressed protein could react with antibodies in sera of SARS patients during convalescence. This provided the basis for the further study on SARS virus vaccine and diagnostic agents.
