猫杯状病毒VP1及其截短体VP1-CDE重组蛋白的表达纯化与免疫效果评价OA

The study aimed to construct and express VP1 and its truncated version VP1-CDE of feline calicivirus(FCV),and evaluate their immunogenicity.The target genes were cloned into the pET28a-SUMO vector and transformed into E.coli BL21(DE3).The soluble expression was induced by IPTG.The molecular weights of VP1 protein and VP1-CDE protein were about 60 kDa and 15 kDa,respectively,which were consistent with the theoretical values.This was confirmed by Western-blot assay with 6 ×His-tagged antibody.The recombinant proteins were purified by Ni2+affinity chromatography and emulsified with the same volume of Freund adjuvant.BALB/c mice were immunized with recombinant protein VP1 and VP1-CDE.Serum samples were collected for indirect ELISA.The results showed that both the VP1 and VP1-CDE groups induced high titers of specific IgG against the corresponding proteins,and the antibody levels increased with the number of immunizations during the first three immunizations.There was no significant difference in the antibody level 14 days after the fourth immunization and 14 days after the third immunization,and the antibody level reached a plateau.The neutralization test showed that the neutralizing antibody titer of VP1-CDE group was 1∶223,which was similar to 1∶178 of VP1 group 14 days after the third immunization.In summary,VP 1-CDE achieves similar immunogenicity to full-length VP1 with a smaller molecular weight,and is easier to achieve correct folding and less to form inclusion bodies during expression.Therefore,VP1-CDE can be used as the preferred antigen for FCV subunit vaccine,which provides experimental basis for subsequent vaccine development.