猪丹毒丝菌SpaA基因原核表达及表达蛋白质的免疫原性分析

doi:10.11843/j.issn.0366-6964.2017.03.012

Abstract

Abstract:

The aim of this study was to clone and express the SpaA gene of Erysipelothrix rhusiopathiae, and analyze its immunogenicity, thus provide the basis for the development of a new swine erysipelas vaccine. The SpaA gene was amplified using PCR and then connected to the pGEX-6P-1 vector. The positive recombinant plasmid was transformed into Rosetta (DE3) of E. coli and SpaA protein was expressed by induction of IPTG. The expression product was analyzed using SDS-PAGE. The fusion protein was purified by GST affinity chromatography and identified by western blot. The mice were immunized with purified SpaA protein, and then the IgG antibody and cytokine levels of Th1, Th2 were measured with an indirect ELISA. All immunized mice were challenged by Erysipelothrix rhusiopathiae with different pathogenicity, and the pathological changes and the protective rate were observed. The results indicated that the SpaA gene of Erysipelothrix rhusiopathiae was successfully expressed in E. coli, and the molecular weight of purified target protein was 75 kD, which can specific recognize antiserum of Erysipelothrix rhusiopathiae. SpaA can induce high levels of specific IgG antibody in mice, and the levels of TNF-β, IFN-γ, IL-5, IL-10 were significantly increased. The immune challenge-protection rate was 100%. There were obvious differences between pathological changes in control group and experimental group. The experiment suggests that SpaA recombinant protein has good antigenicity, which can stimulate the humoral and celluar immunity, and produce powerful immune protection. Therefore, it can be used as a candidate antigen for developing subunit vaccine of swine erysipelas.

Key words: Erysipelothrix rhusiopathiae, SpaA gene, prokaryotic expression, immunogenicity, immune protection

CLC Number:

S855.12

YAO Yan-bin，LU Ping， YANG Zhi-peng， WEI Jian-zhong，SUN Pei，LI Yu. Prokaryotic Expression and Immunogenicity Analysis of SpaA Gene in Erysipelothrix rhusiopathiae[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2017, 48(3): 492-500.

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Prokaryotic Expression and Immunogenicity Analysis of SpaA Gene in Erysipelothrix rhusiopathiae

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