日本血吸虫鞘脂激活蛋白样蛋白的基因克隆表达与免疫原性分析  

Cloning,expression and immunogenicity analysis of saposin-like proteins of Schistosoma japonicum

作　　者：张纯[1] 戴洋[1] 王晓婷[1] 鲁飞[1] 唐建霞 赵松[1] 朱荫昌[1] 

机构地区：[1]江苏省血吸虫病防治研究所、卫生部寄生虫病预防和控制技术重点实验室、江苏省寄生虫分子生物学重点实验室,无锡214064

出　　处：《中国血吸虫病防治杂志》2010年第3期228-232,共5页Chinese Journal of Schistosomiasis Control

摘　　要：目的克隆并表达日本血吸虫鞘脂激活蛋白样蛋白(SLP),并对其免疫原性进行分析。方法根据华支睾吸虫鞘脂激活蛋白样蛋白序列,采用tblastn法比对,在基因库中找出与之同源的日本血吸虫EST片段,并经过DNAStar、Sig-nalP3.0分析获得Sj-SLP基因序列,经PCR扩增从日本血吸虫cDNA中获得的目的基因克隆入原核表达载体pET15b,转化至大肠埃希菌BL21,异丙基巯基半乳糖诱导表达并纯化;采用Westernblot分析其抗原性;用纯化的SLP重组蛋白免疫兔,ELISA检测免疫原性。结果 PCR扩增获得Sj-SLP基因,约296bp,并成功地进行了原核表达,纯化的表达产物重组Sj-SLP(rSj-SLP)能被感染兔血清识别,而不能被正常兔血清识别。rSj-SLP免疫兔可产生1∶12800的特异性抗体。结论日本血吸虫存在鞘脂激活蛋白样蛋白,rSj-SLP具有较强的免疫原性和抗原性。Objective To clone and express the saposin-like protein of Schistosoma japonicum and analyze its immunogenicity.Methods The homologous EST fragment of S.japonicum with the sequence of saposin-like protein（SLP）of Clonorchis sinensis was found from the genebank by using the tblastn technique,and then the gene sequences of Sj-SLP were obtained by analysis of DNAStar,SignalP 3.0.The gene of Sj-SLP was amplified by PCR from cDNA of S.japonicum.The gene was cloned into plasmid pET15b and expressed in E.coli BL21.The expressed product was purified.Two rabbits were immunized with the purified Sj-SLP,and the anti Sj-SLP antibody in immunized rabbits＇sera was tested with ELISA assay.Results The Sj-SLP gene（about 296 bp）was successfully amplified,cloned and expressed in E.coli BL21.The purified rSj-SLP could be recognized by the infected rabbits＇sera of S.japonicum,but not by healthy rabbits＇sera.The special anti-Sj-SLP antibody titer in immunized rabbits＇sera was high（1∶12 800）.Conclusion It is confirmed that saposin-like protein exists in S.japonicum.rSj-SLP possesses a high aimmunogenicity.

关 键 词：日本血吸虫 鞘脂激活蛋白样蛋白 基因 抗原性 

分 类 号：R973.2[医药卫生—药品]