鹅细小病毒VP3亚单位疫苗的制备及其免疫效果  

Preparation and Immune Effect of VP3 Subunit Vaccine of Goose Parvovirus

作　　者：杨焕章[1] 王凯[1] 陈绍红[2] 刘艳芬[1] 刘铀[2] 

机构地区：[1]广东海洋大学农学院,广东湛江524088 [2]广东海洋大学生化中心,广东湛江524088

出　　处：《动物医学进展》2013年第10期41-45,共5页Progress In Veterinary Medicine

基　　金：广东省科技计划项目(2007A020200006-8)

摘　　要：为了研究鹅细小病毒(GPV)VP3亚单位疫苗的免疫效果,从ZJ-04分离株基因组DNA扩增VP1和VP3基因并克隆到pMD18-T载体,测定其核苷酸序列;将VP3基因亚克隆至质粒载体pET-32a(+),重组质粒pET-VP3用于转化E.coli BL-21(DE3)感受态细胞获得重组菌BL-VP3;以VP3重组蛋白和BL-VP3重组菌制备油乳剂疫苗免疫接种种鹅,用琼脂扩散试验检测血清GPV抗体,观察母源抗体对雏鹅GPV感染的免疫保护效果。结果表明,ZJ-04株VP1基因大小为2 199bp,编码732个氨基酸,与GenBank收录的GPV毒株的核苷酸序列同源性为95.3%～99.2%,氨基酸序列同源性为97.7%～99.3%;SDS-PAGE、Western blot检测结果表明,重组VP3分子质量约80ku,能与GPV抗体发生特异性反应;VP3包涵体和BL-VP3重组菌均能诱导种鹅产生GPV抗体,新生雏鹅可获得良好的免疫保护。The present work was aimed to investigate the immune efficiency of the recombinant subunit vac- cine of goose parvovirus（GPV）. Both of VP1 and VP3 genes were amplified from genome DNA of ZJ-04 strain, the amplified fragments were cloned into pMD-18T vector and sequenced subsequently. The VP3 gene was then subcloned into the expression plasmid pET-32a（q-）, and the recombinant plasmid pET-VP3 was used to transform Ecoli BL-21（DE3） competent cells and the recombinant bacteria BL-VP3 was ob- tained. 20 28-week-old breeding Shitou geese were inoculated with oil emulsion vaccine prepared with re- combinant VP3 inclusion or BL-VP3 respectively, and boosted 2 weeks later. The serum GPV antibody ti- ters of the birds were detected by agar diffusion test, and the mortality of the goslings challenged with ZJ- 04 was observed to evaluate the immune protection efficiency of GPV maternal antibody. The results re- vealed that the VP1 gene of ZJ-04 was consisted of 2 199 bp, encoding for 732 amino acids, and the nucleo- tide sequence homology among ZJ-04 and other reference GPV strains recorded in Genbank were 95.3 %- 99.2%, and the homology of the deduced amino acid sequence also came to 97.7%-99.3%. SDS-PAGE and Western-blot detection showed that the recombinant VP3, with molecular weight of 80 ku, could specifically reacted with GPV antisera. The immunized geese inoculated with VP3 inclusion or BL-VP3 vaccine showed higher serum antibody titers against GPV, and the goslings originated from the immunized geese above showed perfect resisitant to the challenge of ZJ-04.

关 键 词：鹅细小病毒（GPV） VP1基因 VP3基因 亚单位疫苗 

分 类 号：S852.659.2[农业科学—基础兽医学]