日本血吸虫肌钙蛋白T(SjTnT)基因克隆表达及免疫保护效果评估  

Cloning,expression and immuno-protection analysis of a gene encoding troponin T of Schistosoma japonicum(SjTnT)

作　　者：王馨茁 洪炀[2] 韩宏晓[2] 李长健[1,2] 曹晓丹[2] 陆珂[2] 李浩[2] 朱传刚[2] 傅志强[2] 林矫矫[2] 

机构地区：[1]上海师范大学生命与环境科学学院,上海200234 [2]中国农业科学院上海兽医研究所

出　　处：《中国血吸虫病防治杂志》2014年第4期394-398,共5页Chinese Journal of Schistosomiasis Control

基　　金：中国博士后科学基金(2012M510630);上海科技发展基金(12140902700);中国农业科学院院长基金(2013JB18)

摘　　要：目的克隆、表达日本血吸虫肌钙蛋白T(SjTnT)编码cDNA,评估重组抗原抗血吸虫感染的免疫保护效果。方法利用PCR技术扩增SjTnT基因,构建重组表达质粒pET28a(+)-SjTnT并诱导表达,用重组蛋白rSjTnT免疫BALB/c小鼠制备免疫血清。利用免疫印迹试验(Western blotting)和酶联免疫吸附试验(ELISA)检测rSjTnT诱导的免疫反应。采用rSjTnT免疫小鼠,评估其诱导的免疫保护效果。结果成功克隆、表达了日本血吸虫SjTnT基因。Western blotting显示rSjTnT能被该重组蛋白免疫小鼠血清识别,具有良好的免疫原性。rSjTnT能诱导小鼠产生高水平的特异性IgG抗体。动物免疫保护试验表明,rSjTnT能诱导小鼠产生33.89%的减虫率以及43.94%的肝脏减卵率。结论制备的rSjTnT在小鼠体内能诱导产生部分抗血吸虫感染的免疫保护效果,为评估SjTnT的生物学功能奠定了基础。Objective To clone cDNA encoding troponin T of Schistosoma japonicum（SjTnT）,and evaluate the protective efficacy induced by recombinant SjTnT in BALB/c mice against S. japonicum challenge infection. Methods The SjTnT gene was amplified from 28-day-schistosome cDNAs by PCR and then subcloned into pET28a（＋）. The recombinant SjTnT protein （rSjTnT）was expressed in Escherichia coli BL21（DE3）cells. The serum specific to rSjTnT was prepared by immunized BALB/c mice with the recombinant antigen,and the immunogenicity of rSjTnT was detected by Western blotting and ELISA. The immuno-protective efficacy induced by rSjTnT in BALB/c mice was evaluated according to the reduction in worm and egg counts. Results The cDNA encoding SjTnT was successfully cloned and expressed in E. coli. Western blotting showed that rSjTnT had a good immunogenicity. The high level of specific IgG antibodies was detected,and 33.89% worm reduction and 43.94% liver egg reduction were obtained in mice vaccinated with rSjTnT combined with Seppic 206 adjuvant compared with those in the adjuvant control group. Conclusions rSjTnT could induce partial immuno-protection against S. japonicum infec-tion in BALB/c mice. This study provided a basic for understanding the biological function of SjTnT.

关 键 词：日本血吸虫 肌钙蛋白T 克隆 表达 免疫保护 效果评估 

分 类 号：R383.24[医药卫生—医学寄生虫学]