机构地区: 广东省农业科学院
出 处: 《华南农业大学学报》 2011年第4期53-56,共4页
摘 要: 利用PCR方法从广东番茄黄化曲叶病毒分离物G3(Tomato yellow leaf curl Guang dong virus-[G3],TYLCGuV-[G3])全长序列的重组质粒上扩增该病毒的AC2基因,将其构建至原核表达载体pET-28b(+)上,经IPTG诱导,在大肠埃希菌Rosetta(DE3)Ⅲ中高效表达.SDS-PAGE电泳结果显示,目的融合蛋白与预期大小一致,相对分子质量约为19 500.以诱导的融合蛋白为抗原,免疫注射新西兰大耳白兔制备抗血清,间接ELISA测定抗血清效价为1∶16 384.Western-blotting检测结果表明,制备的抗血清有较好的抗原-抗体识别反应,为专化性抗血清. AC2 gene of Tomato yellow leaf curl Guangdong virus isolate G3 ( TYLCGuV- [ G3 ] ) was amplified from its recombinant plasmid by PCR, and the gene was cloned into prokaryote expression vector pET-28b( + ). The expressed fusion protein was highly expressed in Escherichia coli Rosetta( DE3 )Ⅲ induced by IPTG, and the molecular mass was about 19 500 by SDS-PAGE analysis. The fusion protein was used as an antigen to immunize the healthy rabbit and the antiserum of AC2 was prepared. The titer measured by ELISA was about 1:16 384. Western-blotting analysis indicated that the antiserum could serologically react with AC2 protein of TYLCGuV-[ G3 ].
关 键 词: 广东番茄黄化曲叶病毒 基因 原核表达 抗血清制备