Cloning And Expressio Omp2b Gene For Brucella.ovis (80/019) Strain

In this paper,a pair of primers were designed according to the nucleotide sequence of omp2b gene of Brucella ovis reported。With the specific primers,one target fragment about1.1kb was amplified from the genomic DNA via PCR of the Brucella ovis 80/019 strain isolated in XingJiang。The target fragment was inserted into the linearized plasmid vector pBS-T。The recombinant plasmid was proved to be true by enzyme analysis and PCR identify。In order to further identify the credibility of the gene,the sequence was obtained by sanger,s sequencing technique。Then Compared the omp2b of XingJiang Brucella ovis with the reported omp2b gene sequence of Brucella ovis and Brucella melitensis which shared 89.72% homology with Brucella ovis and shared 96.10% homology with Brucella melitensis。Phylogenetic tree on structural gene omp2b obtained from Brucella ovis of XingJiang 80/019 and other omp2b gene showed the gene we cloned is so near related Brucella melitensis。To express omp2b antigen protein from out membrane protein of Brucella ovis in XinJiang,study the probability for it as subunit vaccine and diagnose antigen。The fragment was identifeied and cloned into prokaryotic expression vector pET--28a(+)。At last,the combinant plasmid was transformed into E.coli BL21(DE3)and induced with IPTG to expressen coded proteins。By analysis of SDS-PAGE and Western-blot。An expression band about 38kDa protein was found。we obtained omp2b gene fragment and expressed successfully in Escherichia coli。...