猪痘病毒（SWPV）具有基因组大、复制非必需区（NER)多、宿主特异性严格、可插入大片段外源基因等特点，使其特别适合作为疫苗载体。但SWPV作为疫苗载体的关键问题有待解决，如合适NER的筛选、不同NER对外源基因表达及免疫学特性的影响、强启动子的筛选、毒力基因的鉴定等。本项目拟以SWPV江西株为亲本病毒，采用融合PCR、线性DNA同源重组等技术，构建一系列的NER缺失毒株，并分别测定其生物学特性；在此基础上，选择不同的NER分别插入PCV2 ORF2基因，构建表达PCV2衣壳蛋白的重组SWPV，并分别测定其外源蛋白的表达及免疫学特性。本项目将筛选、鉴定出SWPV的一系列NER，解析不同NER的生物学功能；通过系统分析不同NER对插入外源基因的体内外表达及免疫学特性的差异，优选出稳定、高效表达外源基因的NER；分析不同NER与SWPV毒力的相关性，以期发现适合构建SWPV活载体疫苗的NER。

Swinepox virus (SWPV) has a potential to become an excellent vaccine vector. The SWPV genome is large (146kb) and contains multiple non-essential region (NER), allowing the insertion of many large-sized foreign genes. Despite these favorable characteristics, there exist several major challenges in generating a SWPV-based vaccine vector. These challenges include the difficulties in the identification of virulence genes, identification of foreign gene insertion sites, and screening of strong promoters. Such a vaccine vector is also required to express high expression level proteins with strong immunogenicity. In this project, many genetically engineered vector candidates will be generated by mutating NER of the Jiangxi isolate genome using double-joint PCR and homologous recombination, and then the biological characteristics of mutant SWPV will be analyzed. Optimal foreign gene insertion sites will be identified by screening and comparing different NERs based on a variety of biological characteristics. To evaluate the engineered vectors in vitro and in vivo, we will generate recombinant SWPV strains expressing capsid protein of Porcine circovirus type 2 (PCV2). Pigs will be immunized, followed by comparative analysis of gene expression and immunogenicity levels of recombinant capsid protein. This project is expected to obtain a series of NERs for construction of ideal SWPV-based live vaccine vectors that will significantly promote effective vaccine development in future.