猪口蹄疫O型合成肽疫苗抗原多肽的串联表达及其活性鉴定.pdfVIP

DOI牶牨牥牣牨牭牴牫牫牤j牣cnki牣牨牥牥牬牠牫牪牰牳牣牪牥牨牨牣牨牥牣牥牬牥 　河南农业科学,2011,40(10):126-130 　Journal of Henan Agricultural Sciences O 卢清侠, 郭军庆, 郝慧芳, 杨苏珍, 邢广旭, 杨继飞, 王世红, 张改平* ( /, 450002) :为获得含合成肽疫苗抗原多肽的重组表达蛋白, 根据猪口蹄疫O 型合成肽疫苗中抗原多肽 的氨基酸序列, 用大肠杆菌偏好性密码子人工合成该抗原多肽的双拷贝基因序列,并克隆至表达 载体pET32a 中, 构建原核表达质粒pET32a- 2。将重组质粒转化BL21(DE3), 经I TG 诱导表 达, 收集诱导的菌液进行SDS- AGE 电泳和Western-blot 分析。结果显示,在分子量27kD 处有1 条明显的蛋白表达条带, 且能被O 型FMDV 阳性血清识别。对表达产物进行可溶性分析, 结果显 示,表达蛋白几乎全部以可溶形式存在。表达蛋白经N-i NTA 树脂亲和纯化, SDS- AGE 显示纯 化的目的蛋白在 27kD 处有单 一目的条带,具有较高的纯度。ELISA 检测结果显示, 纯化的重组 蛋白具有较高的活性。结果表明, 该抗原多肽的双拷贝串联重复序列在大肠杆菌中得到了可溶性 高效表达, 为开发诊断试剂和基因工程疫苗奠定了基础。 :猪口蹄疫;合成肽疫苗;抗原多肽;串联表达;蛋白纯化;活性鉴定 :S852. 65　　:A 　　:1004-3268(2011)10-0126-05 Tandem Expression and Activity Analysis of Antigen eptide of Swine Foot and Mouth Disease (Type O)Synthetic eptide Vaccine LU Qing-xia,G UO Jun-qing, HAO Hu-i fang, YANG Su-zhen, * XING Guang-xu, YANG J-i fei,WANG Sh-i hong,ZHANG Ga-i ping (Key Laboratory of Animal Immunology of the Ministry of Agriculture /Henan rovincial Key Laboratory of Animal Immunology, H enan Academy of Agricultural Sciences,Zhengzhou 450002, China) Abstract:In order to get the recombination protein which contained antigen peptide of synthetic peptide vaccine,two-copy gene was synthesized using E . co li optimal codons according to the am-i no acid sequence of antigen peptide. The recombinant expression vector pET32a- 2 was construc- ted by cloning two-copy gene of antigen peptide into the prokaryotic expression plasmid pET32a. The BL21(DE3)was transformed with pET32a-2, and the protein corresponding to 2 gene was expressed after induction with I TG. SDS- AGE and Western-blot with the product of bacterial culture induction with I TG showed