Cloning And Prokaryotic Exppression Of Outer-Membrane Protein Omp31 Gene Of Brucella In Deer

Brucellosis is a zoonotic infectious disease caused by brucella, for which the main host is sheep, cattle and pigs, now the number of deer farms infected with brucella is increasing. Brucella is worldwide circulating and has improved circulation in China in recent years that seriously damage the development of animal husbandry and human health. On present, vaccination for susceptible animals and culling of diseased animals are effective measure to prevent and control brucellosis. Thus, it is vital to research effective and safe vaccine with modern biological techniques as well as test method to detect brucella.The omp31 as Brucella immune protective antigen can induce the production of neutralizing antibodies in a large number of outer membrane protein that having a protective effect. Brucella is an ideal candidate genes in the gene engineering vaccine, and is also used for preferred genes and proteins for clinical diagnosis of brucellosis.In addition, the outer membrane protein Omp31 is highly conserved gene of Brucella virulence. The omp31 gene deletion can reduce the Brucella virulence, so can by deletion omp31 gene to the development of gene deleted vaccine. In this study, the plasmid pMD 18T-Omp31 as template by BamH I and Xho I double digestion, the Omp31 were obtained and subcloned to pET28a prokaryotic expression vector construct pET28a-Omp31 prokaryotic expression vector, which was transformed into E.coli BL21 competent, induced by IPTG, so Omp31 gene expression in E.coli BL21 competent and the recombinant protein were analysed by Western Blot.The results show that the study successfully constructed prokaryotic expression vector pET28a-Omp31, induced by IPTG, which can be stably expressed in E.coli BL21 competent in,Omp31 recombinant proteins in inclusion bodies.the higher the concentration of nickel column purification of the protein, Western Blot test showed its have a good reactionogenicity.This study successfully expressed and purified recombinant protein Omp31 not only higher concentration, but also has good immunogenicity, which laid the foundation for the further development of Brucella subunit vaccine and diagnostic ELISA kits.