摘要
在大肠杆菌中高效表达日本血吸虫 (Schistosomajaponicum ,Sj)Sj -Ts1融合蛋白并测定其免疫保护效果。将Sj-Ts1基因亚克隆至 pGEX - 5X - 3原核载体 ,转化入大肠杆菌ER2 5 6 6 ,并用IPTG对该重组菌进行诱导表达中。将此表达产物进行WesternBlot分析后免疫小鼠 ,免疫剂量为 10 0 μg/次 /鼠。并设蛋白佐剂对照和PBS对照。免疫三次后进行攻击感染 ,计数虫负荷及肝卵负荷。在IPTG诱导下 ,亚克隆至 pGEX - 5X - 3的Sj-Ts1基因在大肠杆菌内高效表达融合蛋白SjGST -Ts1,用此融合蛋白免疫小鼠 ,能诱导产生 2 4 16 %的减虫率和 4 8 74 %的减卵率。Sj-Ts1基因亚克隆至 pGEX -5X - 3载体后可在大肠杆菌中高效表达 ,表达产物可诱导小鼠产生一定程度的抗Sj保护性免疫力。
To subclone and express the Schistosoma japonicum Sj-Ts1 geng in E.coli and to evaluate the immune protective effect of the recombinant proteins,the cDNA of the Sj-Ts1 gene was subcloned to expression vector pGEX-5X-3 and transformed to E.coli ER2566.After induction with IPTG,the expressed fusion proteins were identified by Western blotting,emulsified with Freund's complete adjuvant (FCA) and then immunized mice subcutaneously,with a dosage of 100 μg each time at 0,2 and 6 week,meanwhile,FCA and PBS controls were set up.Mice were challenged with cercariae two weeks after the last immunization,and were perfused 6 weeks after challenge to obtain worm burden and the number of eggs per gram of liver (LEPG).It was found that the Sj-Ts1 gene was subcloned successfully.The fusion protein SjGST-Ts1 was highly expressed in E.coli ER2566 through IPTG induction,and could be identified by immune sera of Sj.The worm reduction rate and the egg reduction rate in the immunized group of mice were 24.16% and 48.74% respectively.It concludes that the Sj-Ts1 gene of Schistosoma japonicum can be expressed in E.coli after subcloning to expression vector pGEX-5X-3 and the expressed fusion protein SjGST-Ts1 can induce protective immunity against Schistosoma japonicum.
出处
《中国人兽共患病杂志》
CSCD
北大核心
2004年第7期592-595,共4页
Chinese Journal of Zoonoses
基金
WHO/TDR资助项目 ( 980 2 68)
湖南省科技厅资助项目( 0 0jzy2 115 )
