BVDV/IBRV主要抗原表位E2/gD基因串联表达及免疫原性

Tandem Expression and Immunogenicity of the Major Antigen Epitopes of BVDV/IBRV E2/gD Gene

为开发研制牛病毒性腹泻-黏膜病毒(BVDV)及牛传染性鼻气管炎病毒(IBRV)的联合亚单位疫苗,开展了BVDV E2蛋白、IBRV gD蛋白主要抗原表位串联表达及免疫原性研究。利用2步PCR法获得E2-gD基因,构建原核表达载体pET28a-E2-gD,转化大肠杆菌BL21感受态细胞,IPTG诱导蛋白表达,SDS-PAGE结果显示重组蛋白表达,BCA工作盒测定纯化蛋白浓度为2.69 mg/mL;Western-blot结果显示其具有作为免疫原的特异性;免疫家兔制备抗血清,血清中和试验结果表明,中和抗体效价为44.7(IBRV)/22(BVDV)。这说明E2-gD蛋白免疫原性较好,可诱导产生较高效价的中和抗体。

In order to study the combined subunit vaccine of bovine viral diarrhea virus （BVDV）and in-fectious bovine rhinotracheitis virus （IBRV）,the experiments which study the immunogenicity of tandem expression of major antigen epitopes of BVDV/IBRV E2-gD gene were carried out .Gene segments were obtained by two-step PCR method,then prokaryotic expression vector pET28a-E2-gD was constructed and transformed into the E .coli competent cells BL21 （DE3）.The cells were induced by IPTG to express protein E2-gD .SDS-PAGE results showed that the recombinant protein was expressed .After purifica-tion,protein concentration was determined to be 2.69 mg/mL by BCA Protein Assay Kit .Western-blot results showed that it had a specific immunogenicity .The protein was used to prepare the antiserum by immunizing rabbits .Antibody neutralization titer of the antiserum was 44.7（IBRV）/22（BVDV）by serum neutralization test .The results showed that E2-gD protein had a better immunogenicity and could induce high titer of neutralizing antibodies .Above all,the research laid a foundation for further study on subunit vaccine which can be used to resist BVDV/IBRV infection .