The Cloning And Prokaryotic Expression Of Truncated GB And GE Genes Of Infectious Bovine Rhinotracheitis Virus

IBRV is the pathogen of infectious bovine rhinotracheitis.This virus genome encodes 12 glycoproteins,including glycoprotein B and glycoprotein E,which is an important protein on the surface of the virus.The gB gene is necessary for virus replication,the gE gene is not.In this study,bioinformatic analysis of the proteins encoded by gB and gE genes of IBRV was carried out and We cloned and expressed the antigen region fragment according to the analysis results.The results show:(1)The IBRV gB protein encodes 945aa,exists in the transmembrane region at 780-802 and 809-831,and has no signal peptide site.The protein chain is in 300-450 position,hydrophilic,antigenicity and high surface accessibility.IBRV gE encodes 597aa,transmembrane region exist in the 422-444 position,and prediction of the presence of signal peptide sites in position 28-29,protein chains in the 110-225 and 450-550 of hydrophilicity,antigenicity,surface accessibility is high.(2)We amplified the truncated fragments of the gB and gE genes with lengths of 279bp and 285bp,respectively.And successfully constructed recombinant plasmids pET-gB and pET-gE,the optimal induction time of recombinant plasmid was 3 hours,and the optimum concentration of IPTG was 1mmol/L.(3)We induced pET-gB and pET-gE proteins in the host expressing bacterium BL21 and obtained proteins about 34KDa in size.gB recombinant protein was expressed mainly in the form of inclusion body,and the gE recombinant protein mainly in soluble form.Western-bolt assay showed that the fusion protein was successfully expressed in E.coli and purified by affinity chromatography with nickel column for gE recombinant protein.This study provided a basis for the establishment of ELISA and for the development of gene deletion vaccines.