Prokaryotic Soluble Expression and Immunogenicity Analysis of VP1 Gene of Foot-and-mouth disease virus serotypes SAT2
LI Guo-Xiu1,3,*, OU Yun-Wen1,2,*, DING Yao-Zhong1, LI Qian1, DAI Jun-Fei1, LIU Lei3,**, ZHANG Jie1,**
1 Office International Des Epizooties (OIE)/National Foot-and-Mouth Disease (FMD) Reference Laboratory/State Key Laboratory of Veterinary Etiological Biology, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, China; 2 Animal Disease Prevention and Control Center of Kaijiang County, Dazhou 636250, China; 3 College of Veterinary Medicine, Gansu Agricultural University, Lanzhou, 730070, China
Abstract：Foot-and-mouth disease serotypes Southern African Territories 2 (SAT2-FMD) is an acute, heat, and highly contactive infectious disease caused by Foot-and-mouth disease virus serotypes Southern African Territories 2 (SAT2-FMDV). At present, global trade and mutual visits are frequent, and the risk of SAT2-FMD entering China is extremely high, which will seriously threaten the sustainable and healthy development of Chinese breading industry. This experiment was aimed to prokaryotic soluble expression and analysis the immunogenicity of VP1 gene of SAT2-FMDV (GenBank No. JX014256). The gene was amplified from the recombined plasmid pUC57-His-SUMO-SAT2-VP1 by PCR, with a product of approximately 993 bp. The identified recombined plasmid pET32a-HisSUMO-VP1 was expressed by Escherichia coli BL21 (DE3) and was induced by 1 mmol/L isopropy-β-D-thiogalactoside (IPTG). The HisSUMO-VP1 protein was purified by Ni-NTA, and the purified protein was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDA-PAGE) and Western blot. The HisSUMO-VP1 protein was used as an immunogen for immunization of Balb/C mice (Mus musculus), and the serum antibody, spleen lymphocyte proliferation and cytokine levels were detected. The results of SDA-PAGE and Western blot showed that the HisSUMO-VP1 protein with a relative molecular weight of 63 kD was highly soluble and had good reactivity. The immune experiment found that the serum antibody levels in the immunized mice vaccinated with the HisSUMO-VP1 fusion protein group was significantly higher than those in the HisSUMO protein vaccinated group and the mock group (P<0.001), and the mice spleen lymphocytes had a significant lymphocyte proliferation response (P<0.001). The levels of IFN-γ and IL-2 were significantly higher than those in the PBS group (P<0.05). The HisSUMO-VP1 protein with excellent immunogenicity is solublely expressed in E. coli, providing data support for the diagnosis and vaccine development of SAT2-FMDV.
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