摘要
应用PCR技术,以含有新城疫病毒F48E8株全长融合蛋白(F)基因的真核表达质粒pVAX1-F为模板,扩增出594bp大小的F基因的部分片段。经克隆筛选和测序后,构建成重组原核表达质粒pGEX-6P-1-F。重组质粒转化表达菌BL21,获得重组菌BL21(pGEX-6P-1-F)。经IPTG诱导和SDS-PAGE分析表明,F基因在原核表达体系中获得高效表达,表达量占菌体总蛋白的23%。Western blotting结果显示,在相对分子质量46ku位置有特异性条带。动物实验结果表明,F蛋白免疫鸡产生的针对新城疫病毒F蛋白的血清抗体效价显著高于空白对照组(P<0.05),说明原核表达系统表达的F蛋白具有良好的免疫原性。
A part of the fusion protein （F） gene which is 594 bp of Newcastle disease virus was amplified by polymerase chain reaction （PCR） from /he recombinant eukaryotic expression plasmid pVAX1-F, and subcloned into prokaryotic expression vector pGEX-6P-1. The F gene was identified by sequencing. Then the recombinant plasmid was transformed into E.coli BL21, and the recombinant was designated as BL21 （pGEX-6P-1-F）. The result of SDS-PAGE verified that there was a desired recombinant protein with molecular weight of 46 000 and accounted for 23% of the total bacterial protein. Western blotting analysis further indicated this expressed product had the immunogenicity of F protein： The F protein could significantly elicit specific serum immune response in immunized SPF chickens,compared with blank control （P〈0.05）. These results demonstrated that the F protein expressed by E.coli has good immunogenicity.
出处
《中国家禽》
北大核心
2009年第20期21-24,共4页
China Poultry
基金
国家自然科学基金资助项目(30871860)
高等学校科技创新工程重大项目培育资金项目(706032)
江苏省自然科学基金项目(BK2007511)
江苏省高校自然科学基础研究项目(07KJB230136)
关键词
新城疫病毒
融合蛋白
原核表达
免疫原性
Newcastle disease virus
fusion protein
prokaryotic expression
immunogenicity
