Expression and purification of SARS coronavirus membrane protein

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...

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Detalles Bibliográficos
Autores principales: Wuxing, Dai, Mingjun, Lei, Shaoting, Wu, Zhihao, Chen, Liang, Liang, Hujrong, Pan, Li, Qin, Shitong, Gao, Shishan, Yuan, Renli, Zhang
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Huazhong University of Science and Technology 2004
Materias:
Article
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7089427/
https://www.ncbi.nlm.nih.gov/pubmed/15641679
http://dx.doi.org/10.1007/BF02831095
Descripción
Sumario: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 intoEscherichia coli (E. Coli) BL21 (DE3) and induced by Isopropyl-β-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.

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