弓形虫分泌蛋白ROP2的原核重组表达与免疫反应性鉴定  

RECOMBINANT PROKARYOTIC EXPRESSION AND IMMUNOREACTIVITY ANALYSIS OF THE ROP2 SECRETORY PROTEIN OF TOXOPLASMA GONDII

作　　者：薛峰[1,2] 黄敏君[1,2] 洪彩玲[1] 杨英超[3] 甘绍伯[1,2] 谷俊朝[1,2] 

机构地区：[1]首都医科大学附属北京友谊医院,北京热带医学研究所,北京100050 [2]热带病防治研究北京市重点实验室,北京100050 [3]中国食品药品检定研究院,生物制品检定所,寄生虫疫苗室,北京100050

出　　处：《寄生虫与医学昆虫学报》2014年第2期75-80,共6页Acta Parasitologica et Medica Entomologica Sinica

基　　金：国家自然科学基金项目（No.81071375 to JG,81301404 to FX）;北京市自然科学基金项目（No.7132042to JG,7144196to FX）;北京市优秀人才培养资助D类项目（No.2013D003034000021 to FX）;首都医科大学基础-临床科研合作基金项目（No.13JL16 to FX）;首都医科大学附属北京友谊医院科研启动基金项目（No.2011_30yykyqd to FX）

摘　　要：本研究在大肠杆菌工程菌中重组表达弓形虫棒状体蛋白2（ROP2），并初步评价其免疫反应性。主要步骤为体外细胞培养速殖子，提取总RNA，反转录获得cDNA，PCR扩增ROP2基因全长，与pET-41 Ek／LIC载体通过基因重组直接连接，在大肠杆菌Rosetta^TM（DE3）pLysS工程菌中诱导表达重组融合蛋白GST-6×His—ROP2，最后以重组蛋白与人源弓形虫抗血清的Western blotting反应评价rROP2免疫反应性。本研究获得了ROP2全长重组蛋白，初步证明其良好的免疫反应性，为进一步改进弓形虫免疫学诊断方法，研究分泌蛋白与宿主细胞的相互作用奠定了基础。In this study, we expressed the recombinant protein of the Toxoplasma rhoptry protein ROP2 in E. coli, and primarily evaluated the immunoreactivity of the recombinant protein. The Toxoplasma gortdil tachyzoite were cultivated in vitro in cell line, the cDNA were synthetized from the total RNA. The total length of ROP2 was amplified using PCR, and ligated into the pET-41 Ek/LIC vector by direct nucleotide sequence recombination. The recombinant fusion protein GST-6 × His-ROP2 was induced in E. coli strain Rosetta^TM （DE3） pLysS, and the immunoreactivity of rROP2 was analyzed by Western blotting of the recombinant protein and human anti-T, gondii serum. The full length of ROP2 recombinant protein was obtained and its good immunoreactivity was verified primarily in this study, which set foundation for further improving immunological diagnosis method, and study on protein interaction between T. gondii secretory protein and host ceils.

关 键 词：刚地弓形虫 棒状体蛋白ROP2 原核重组表达 免疫反应性 

分 类 号：Q55[生物学—生物化学]