日本血吸虫肌动蛋白cDNA的克隆表达及其免疫保护功能  

CLONING,EXPRESSING AND PROTECTIE IMMUNITY EVALUATING OF ACTIN cDNA OF SCHISTOSOMA JAPONICUM

作　　者：朱建国[1,2] 林矫矫[1] 冯新港[1] 陈永军[1] 吴祥甫[2] 周元聪[2] 蔡幼民[1] 

机构地区：[1]中国农业科学院上海家畜寄生虫病研究所,农业部动物寄生虫学重点开放实验室,上海200232 [2]中国科学院上海生物化学研究所

出　　处：《中国人兽共患病杂志》2001年第5期48-52,共5页Chinese Journal of Zoonoses

基　　金：国家高技术"863"基金 (No 10 2 -0 7-0 4-0 3)资助项目

摘　　要：根据曼氏血吸虫肌动蛋白cDNASmAct2设计一对引物 ,以日本血吸虫中国大陆株成虫mRNA为模板 ,用RT -PCR法扩增出一大小为 113 1bp的cDNA片段。测序后分析序列推断该片段为编码肌动蛋白基因的完整阅读框cDNA ,与SmAct2cDNA序列同源性为 92 %。将其克隆到表达载体pET2 8a( + )中 ,在大肠杆菌BL2 1中获得了较高量的表达 ,融合表达产物分子量约为 4 3kDa。利用日本血吸虫成虫粗抗原免疫大白兔所得抗血清对该表达产物进行Western印迹检测 ,在预测位置出现了明显的识别条带 ,说明所克隆的cDNA表达产物具有良好的抗原性。用该表达产物免疫昆明系小鼠 ,攻击血吸虫尾蚴以进行保护性实验 ,结果　其减虫率为 2 8 4 % ,减卵率为 2 9 2 % ,初步结果显示血吸虫肌动蛋白具有一定的免疫保护功能。A 1 131bp cDNA fragment was amplified by RT-PCR from adult Schistosoma japonicum(Chinese strain)mRNA with primers designed according to published SmAct2 encoding actin Sequence analysis indicated that this fragment, named SjAct,with 92% homology to SmAct2,was a ORF of cDNA encoding actin of Schistosoma japonicum (Chinese strain) This fragment was cloned into the expression vector pET28a(+) and subsequently expressed in Escerichia coli SDS-PAGE revealed that the molecular weight of this expressed product(rSjAct) was around 43kDa Western Blot showed that the recombinant protein had good antigenicity when probed with the rabbit serum against S japonicum(S j) worm antigens In order to evaluate the protective efficacy induced by this recombinant protein,Kunming mice were vaccinated 3 times with rSjAct and then challenged with 40 cercariae of S j The mice were perfused 7 weeks post-infection The reduction rate of worm and egg count per gram liver in vaccinated mice was 28 4% and 29 2% respectively The preliminary results showed that the rSiAct induces the significantly protection in vaccinated animals

关 键 词：日本血吸虫 肌动蛋白 RT-PCR 克隆 基因表达 免疫保护性 CDNA 

分 类 号：R532.21[医药卫生—内科学] R383.24[医药卫生—临床医学]