猪瘟病毒E2蛋白重组T4噬菌体的构建及免疫学特性

CSFV E2 Gene Display on Bacteriophage T4 and the Immunological Activity of Its Expression Product

利用重组 PCR技术将猪瘟病毒 (CSFV) E2基因与 T4噬菌体 SOC基因 C端融合 ,构建了大小为 12 33bp的SOC/ E2融合基因 ,将其插入携带 T4溶菌酶基因 (e)和 denv基因的 T4重组载体 (p RH) ,构建了重组载体 p RSE2 ,通过重组载体与缺失突变型 T4噬菌体基因发生同源重组 ,将 SOC/ E2融合基因整合入 T4噬菌体的基因组中。经EL ISA、Western blot等免疫学检测证实 ,T4噬菌体表达的 E2融合蛋白具有 CSFV免疫学活性 ,动物试验证实 T4 .SOC.

The T4 phage display is a newly developed system for displaying of exogenous peptides or proteins domains by fusion expression on the T4 surface.In this system the displayed proteins maintain their relatively independent conformation and biological activities.This process is initiated by fusing a foreign gene to the T4 surface capsid accessory protein gene SOC(small outer capsid protein).In the present study,The protective antigenic E2 gene of classical swine fever virus(CSFV),was fused in frame to 3′ end of SOC in display vector.The SOC/E2 fusion genes were then integrated into the SOC site of the genome of SOC-deleted T4 phage mutant by homologous recombination leading to the display of the E2 protein on the surface of T4 phage.In western blot the correct bands corresponding to the molecular weights of fused SOC/E2 proteins were visualized.With a panel of CSFV-specific polyclonal antisera in an ELISA assay demonstrated that the bound peptides were displayed in a form accessible for antibodies.Inoculation of mice with recombinant phage T4.SOC.E2 and T4.SOC.mE2 showd a significant immune response as indicated in ELISA using intact CSF virus.The detectable antibody level persisted for 56 days of a complete immunization trial.The antibody titer elicited by T4.SOC.E2 was higher than that elicited by T4.SOC.mE2 indicating the display of full E2 is more antigenic than partial E2.