旋毛虫成虫抗原基因的原核表达及表达产物的特性分析

中国寄生虫学与寄生虫病杂志 ›› 2003, Vol. 21 ›› Issue (1): 6-19.

旋毛虫成虫抗原基因的原核表达及表达产物的特性分析

诸欣平¹,杨静¹,杨雅平¹,Pascal Boireau²,詹斌³,冯建军³,Peter Hotez³

1 首都医科大学基础医学院寄生虫学教研室,北京 100054 2 法国食品安全局 3 美国乔治华盛顿大学微生物与热带医学系,华盛顿200037

收稿日期:1900-01-01 修回日期:1900-01-01 出版日期:2003-02-28 发布日期:2003-02-28

Prokaryotic Expression and Characterization of an Antigenic Gene of Adult Trichinella spiralis

ZHU Xin-ping¹,YANG Jing¹,YANG Ya-ping¹, Pascal Boireau² ZHAN Bin³ FENG Jian-jun³,Peter Hotez³

1 Department of Parasitology,Capital University of Medical Sciences,Beijing 100054,China, 2 Umr Bipar Inra Afssa Enva Upvm,Maisons Alfort,France, 3 Department of Microbiology & Tropical Medicine,the George Washington University,Washington,DC,20037

Received:1900-01-01 Revised:1900-01-01 Online:2003-02-28 Published:2003-02-28

摘要/Abstract

摘要： 　　目的　获得旋毛虫成虫抗原基因的原核表达产物并对其进行纯化及免疫特性分析。　方法　旋毛虫成虫Ts87基因片段亚克隆入PET-28a(+)表达载体,异丙基硫代-β-D-半乳糖苷(IPTG)诱导表达。亲和层析和电洗脱法纯化表达产物,SDS-PAGE、ELISA和Westernblotting对表达产物进行分析鉴定,并将纯化的表达产物人工免疫兔。结果　SDS-PAGE结果显示,表达产物为分子量约为40kDa的重组蛋白。纯化后免疫兔获得高滴度的抗血清。该基因重组蛋白可被感染旋毛虫的猪、兔血清及人工免疫兔血清所识别,与感染囊尾蚴、棘球蚴的患者血清或感染日本血吸虫的兔血清不发生交叉免疫反应。　结论　获得一个新的旋毛虫成虫Ts87基因重组蛋白,该蛋白具有特异的抗原性。

关键词: 旋毛虫, 基因表达, 纯化, 重组蛋白, 抗原, 鉴定

Abstract: 　Objective To obtain a purified Ts87 gene expression product of adult Trichinella spiralis and identify its immunogenicity. Methods Ts87 cDNA was subcloned into PET-28a(+) expression vector. The transformed bacteria bearing PET-28a(+)/Ts87 plasmids were induced by IPTG for production of fusion proteins. The expression product purified with His binding affinity chromatography and electro elution assay was analyzed by SDS-PAGE, ELISA and Western blot, and was used to immunize the rabbits. Results PET-28a(+)/Ts87 transformed bacteria produced the desired fusion protein with a relative molecular weight of 40 kDa. The antisera with high titer were obtained by immunizing rabbit with Ts87 recombinant protein. Ts87 expression protein was detected as positive reaction with infected rabbit sera, infected swine sera and antisera against Ts87 by ELISA. Ts87 protein was also recognized by above-mentioned sera with Western blotting. However, Ts87 protein failed to react with the patient sera infected with Cysticercus cellulosae or Echinococcus granulosus, and rabbit sera infected with Schistosoma japonicum. Conclusion A new Trichinella recombinant protein with specific antigenicity was obtained.

Key words: Trichinella spiralis, gene expression, purification, recombinant protein, antigen, identification

诸欣平;杨静;杨雅平;PascalBoireau;詹斌;冯建军;PeterHotez. 旋毛虫成虫抗原基因的原核表达及表达产物的特性分析[J]. 中国寄生虫学与寄生虫病杂志, 2003, 21(1): 6-19.

ZHUXin-ping;YANGJing;YANGYa-ping;PascalBoireauZHANBinFENGJian-jun;PeterHotez. Prokaryotic Expression and Characterization of an Antigenic Gene of Adult Trichinella spiralis[J]. Chinese Journal of Parasitology and Parasitic Diseases, 2003, 21(1): 6-19.

[1]	姜文静, 孟雅莉, 赵利娜, 王春苗, 张晓磊. 刚地弓形虫棒状体蛋白18和膜表面抗原30复合核酸疫苗对小鼠的免疫保护作用[J]. 中国寄生虫学与寄生虫病杂志, 2023, 41(5): 532-538.
[2]	张旭, 孙希萌. 旋毛虫感染免疫逃逸机制研究进展[J]. 中国寄生虫学与寄生虫病杂志, 2023, 41(4): 492-496.
[3]	芦星, 王水怡, 陈林军, 刘明明, 刘雨桐, 朱慧茹, 姜冰冰, 杜少磊, 巴音查汗, 刘丹丹, 张伟. 马泰勒虫棒状体颈部蛋白5基因的克隆与原核表达[J]. 中国寄生虫学与寄生虫病杂志, 2023, 41(4): 497-501.
[4]	叶井明, 何威, 刘慧媛, 鱼潇, 罗波, 刘美辰, 周必英. 猪囊尾蚴排泄分泌抗原TPx对仔猪树突状细胞活化的影响[J]. 中国寄生虫学与寄生虫病杂志, 2023, 41(3): 286-293.
[5]	郝会囡, 程永康, 张茹, 韩璐璐, 宋艳艳, 龙绍蓉, 刘若丹, 张玺, 王中全, 崔晶. 旋毛虫新生幼虫可溶性抗原的免疫蛋白组学分析[J]. 中国寄生虫学与寄生虫病杂志, 2023, 41(2): 176-182.
[6]	焦红杰, 齐文静, 郭刚, 包建玲, 吴川川, 宋传龙, 李军, 张文宝, 严媚. 细粒棘球蚴抗原B对小鼠巨噬细胞RAW264.7的极化作用[J]. 中国寄生虫学与寄生虫病杂志, 2023, 41(1): 23-28.
[7]	陈齐鲁, 王旭, 李春阳, 官亚宜. 线粒体基因在带绦虫科分子分类鉴定及系统进化研究中的应用[J]. 中国寄生虫学与寄生虫病杂志, 2022, 40(6): 767-773.
[8]	邹伟浩, 吴蔚玲, 廖远鹏, 陈敏, 彭鸿娟. 刚地弓形虫抗缓殖子期抗原1单克隆抗体的制备与应用[J]. 中国寄生虫学与寄生虫病杂志, 2022, 40(5): 587-593.
[9]	李润花, 殷国荣. 刚地弓形虫病多表位疫苗的研究进展[J]. 中国寄生虫学与寄生虫病杂志, 2022, 40(5): 661-667.
[10]	陈兵, 张国莉, 张高红. 血吸虫病候选疫苗临床研究进展[J]. 中国寄生虫学与寄生虫病杂志, 2022, 40(4): 511-515.
[11]	乔飞, 刘鏊, 余复昌, 齐萌. 1例牛带绦虫感染的诊断和鉴定[J]. 中国寄生虫学与寄生虫病杂志, 2022, 40(3): 285-287.
[12]	于嘉利, 刘蕾, 杨博, 楚瑞林, 孙毅凡, 刘耀宝, 程洋. 重组卵形疟原虫裂殖子表面蛋白1 N端的抗原性及免疫原性分析[J]. 中国寄生虫学与寄生虫病杂志, 2021, 39(6): 746-752.
[13]	刘路瑶, 杨聪山, 章翔, 程蓉蓉, 洪宇航, 吴成龙, 孙萌, 陈木新, 艾琳, 徐前明. 孔雀源异刺线虫的形态学特征及分子鉴定[J]. 中国寄生虫学与寄生虫病杂志, 2021, 39(6): 816-820.
[14]	蔡子涵, 曹颖, 朱逢龙, 李倩, 和艳红, 杨毅梅. 纳米金棒标记技术应用于旋毛虫感染诊断的研究[J]. 中国寄生虫学与寄生虫病杂志, 2021, 39(5): 652-658.
[15]	吕立红, 张金成, 胡永红. 蜱类保护性抗原研究进展[J]. 中国寄生虫学与寄生虫病杂志, 2021, 39(4): 542-547.