日本血吸虫(中国大陆株)Sj16基因的原核表达  

PROKARYOTIC EXPRESSION OF Sj16 GENE OF SCHISTOSOMA JAPONICUM CHINESE STRAIN

作　　者：卞国武[1] 余新炳[1] 吴忠道[1] 徐劲[1] 周俊梅[1] 单志新[1] 马长玲[1] 邵筱[1] 

机构地区：[1]中山医科大学寄生虫学教研室,广州510089

出　　处：《中国血吸虫病防治杂志》2001年第6期333-336,共4页Chinese Journal of Schistosomiasis Control

基　　金：国家自然科学基金 ( No.30 0 0 70 6 83);国家教育部博士点基金 ( No.2 0 0 0 45 );广东省自然科学团队研究项目资助

摘　　要：目的　进一步研究日本血吸虫 (中国大陆株 ) Sj16的免疫调节功能 ,丰富有关血吸虫免疫逃避知识。方法　根据曼氏血吸虫 Sm16基因已知序列设计合成一对引物 ,用 PCR技术从日本血吸虫 (中国大陆株 )成虫 c DNA文库中扩增 Sj16基因 ;将 Sj16基因定向克隆入 p GEX- 4T- 1,转化感受态 BL 2 1/ DE3菌 ;用酶切、PCR扩增鉴定筛选得到的重组阳性克隆。 IPTG诱导 p GEX- 4T- 1- Sj16转化的 BL 2 1/ DE3菌 ,用 SDS- PAGE和 Wester- blot分析表达产物。结果　从日本血吸虫 (中国大陆株 )成虫 c DNA文库中获取 Sj16基因 ,重组质粒中含有 Sj16基因 ,成功构建日本血吸虫 (中国大陆株 ) Sj16基因原核表达重组质粒 p GEX- 4T- 1- Sj16 ,p GEX- 4T- 1- Sj16转化的 BL 2 1/ DE3菌经 IPTG诱导可表达与 GST融合的蛋白 (约 40 k Da) ,抗 GST抗体能特异性识别该蛋白。结论　成功原核表达了 Sj16蛋白 ,为深入研究 Sj16的免疫调节功能 ,丰富有关血吸虫免疫逃避知识打下了基础。Objective To further study the immunodulating function of Schistosoma japonicum Sj16. Methods A couple of primers were designed according to the known sequence of Schistosoma mansoni Sm16 and the Sj16 gene was obtained by amplification from cDNA library of Schistosoma japonicum Chinese strain by using PCR technique. By cloning Sj16 gene into a prokaryotic expression vector, pGEX-4T-1-Sj16, a recombinant pGEX-4T-1-Sj16 was constructed and transferred into E.coli BL21/DE3. The positive recombinant pGEX-4T-1- Sj16 was screened and identified by agarose gel electrophoresis, endonuclease digestion and PCR technique. The BL21/DE3 which was transformed with pGEX-4T-1- Sj16 was induced by IPTG, and the expressed protein was analyzed by SDS-PAGE and Western-blot. Results The specific fragment of Sj16 was amplified and the recombinant prokaryotic expression plasmid pGEX-4T-1- Sj16 was successfully constructed. After being induced by IPTG, the transformed BL21/DE3 expressed an about 40 kDa protein which was fused with GST. This protein could be recognized by anti-GST antibody in Western-blot test. Conclusion The recombination prokaryotic expression plasmid pGEX-4T-1- Sj16 was constructed and the Sj16 gene was successfully expressed in BL21/DE3, which paved the way for further study on immunomodulatory function of Schistosoma japonicum Sj16 and obtained the knowledge of the immune evasion of schistosome.

关 键 词：日本血吸虫 Sj16基因 原核表达 免疫调节功能 

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