关键词: 基因佐剂；基因，SPreS2；真核表达载体；瞬时表达

Abstract:

Objective     To construct and verify a recombinant eukaryotic expression plasmid pVAX1-SPreS2 for the further development of a new genetic adjuvant to enhance or adjust humoral and cellular immune responses to a Toxoplasma gondii DNA vaccine. Methods    The S and PreS2 genes from HBV were fused with a polypeptide linker (GSGSGS) by overlapping extension PCR, constructing the cloning plasmid pMD18T-SPreS2 and recombinant eukaryotic expression plasmid pVAX1-SPreS2. Then the construction was verified by PCR, restriction enzyme digestion and sequencing. The specific expression protein SPreS2 was detected by SDS-PAGE and Western blot after transient transfection of pVAX1-SPreS2 into HFF cells, which were treated with lipofectin to induce the establishment of an in vitro eukaryotic cell transient expression system. Results    PCR, restriction enzyme digestion and sequencing results showed the correct construction of the recombinant cloning plasmid pMD18T-SPreS2 and eukaryotic expression plasmid pVAX1-SPreS2. It was found that the fusion gene SPreS2 was transiently expressed in eukaryotic cells and its product had, to some extent, an immunological activity. Conclusion     The recombinant eukaryotic expression plasmid pVAX1-SPreS2 has been successfully constructed and it may function as a new genetic adjuvant for a T. gondii DNA vaccine.

Key words: Genetic adjuvant; Genes，SPreS2; Eukaryotic expression plasmid; Transient expression