编码日本血吸虫Argonaute 3蛋白全长cDNA的获得及初步研究  

Cloning and preliminary analysis of a full-length cDNA encoding Argonaute 3 from Schistosoma japonicum

作　　者：陈晶[1,2] 郭素霞[1] 杨燕萍[1] 刘金明[1] 林矫矫[1] 李家奎[2] 程国锋[1] 

机构地区：[1]中国农业科学院上海兽医研究所,农业部动物寄生虫学重点开放实验室,上海200241 [2]华中农业大学动物医学院动物医学系

出　　处：《中国血吸虫病防治杂志》2010年第4期310-314,共5页Chinese Journal of Schistosomiasis Control

基　　金：国家自然科学基金(30901068);上海市科学技术委员会项目(10410703400);上海市人才发展基金(2009032);中央级公益性科研院所基本科研业务费项目(2008JB18、2009JB08)

摘　　要：目的获得日本血吸虫Argonaute3(Ago3)蛋白的全长编码cDNA并对其初步分析。方法利用cDNA末端快速扩增技术(RACE)获得日本血吸虫Ago3蛋白全长编码cDNA,利用原核表达系统对此蛋白片段进行重组表达,采用亲和层析法纯化重组蛋白,并免疫动物制备多克隆抗体。利用Western blot技术分析该蛋白在日本血吸虫中的表达。结果采用RACE获得了日本血吸虫Ago3蛋白编码全长cDNA,长度为2772bp;利用原核表达系统获得了该蛋白片段的重组融合蛋白,并制备了多克隆抗体;Western blot分析表明,该抗体与日本血吸虫全虫蛋白在分子量约为100kDa处具有特异性反应。结论获得了编码日本血吸虫Ago3蛋白的全长编码cDNA,并制备了多克隆抗体,为进一步研究该蛋白的功能奠定了基础。Objective To obtain and preliminarily analyze a full-length cDNA encoding Argonaute 3 （Ago 3） in Schistosoma japonicum.Methods The technique of rapid amplification cDNA ends （RACE） was employed to obtain a full-length cDNA encoding Ago 3 from Schistosoma japonicum （SjAgo 3）.The cDNA fragment encoding SjAgo 3 was cloned into a pET28a （＋） vector to produce the recombinant protein.The recombinant protein was subsequently purified by affinity purification.The polyclonal antibodies were raised in the mice immunized with the purified protein.Results The full-length cDNA of SjAgo 3 was 2 772 bp in length.Bioinformatical analysis indicated that SjAgo 3 contained a PAZ and a PIWI domain,which were the typical characterization for Argonaute protein family.Western blot analysis indicated that the polyclonal antibodies were able to specifically recognize the protein with molecular weight around 100 kDa from lysates of Schistosoma japonicum.Conclusion A full-length cDNA encoding SjAgo 3 is obtained,which provides preliminary information for further investigation of Ago 3 functions in Schistosoma japonicum.

关 键 词：日本血吸虫 Argonaute蛋白 CDNA末端快速扩增技术 非编码小RNA 

分 类 号：R383.24[医药卫生—医学寄生虫学]