Construction and prokaryotic expression of the fusion protein gene encoding MDV gB CTL epitope and chicken HSP108.

[Objective] The study was to supply the basis on developing a new type of MD vaccine. [Method] HSP108 gene was amplified by RT-PCR from SPF chicken liver,and MDV gB CTL epitope amplified by PCR.The fusion gene for expressing gB CTL epitope and chicken HSP108 was obtained by fusing the gene fragment encoding a MDV gB CTL epitope to 5'end of chicken HSP108 gene.And the fusion gene was inserted into pET-28a to construct the recombinant plasmid,named pET-gBHSP108.The recombinant protein named rgBHSP108 was analyzed by SDS-PAGE,solubilized in urea (8 mol/L),purified by His·Bind affinity chromatography and identified by western blotting. [Result] The chicken HSP108,a fragment of about 2.4 kb in length and MDV gB CTL epitope,a fragment of about 330bp were successfully obtained.The cloned HSP108 gene showed 99.7% homology with the HSP108 gene of chicken oviduct (GenBank Accession No.NM 178423),and 99.0% homology with the HSP108 gene of chicken liver (GenBank Accession No. AF 387865) at nucleotide levels.The fusion gene rgBHSP108 was successfully constructed,and rgBHSP108 was expressed as inclusion body and successfully purified. Immunoblotting analysis showed that the expressed fusion protein was recognized by antibody against MDV gB CTL epitope. [Conclusion] This study successfully constructed a fusion gene encoding chicken HSP108 gene and gB CTL epitope gene of MDV.And the fusion gene was successfully expressed by E.coli.