日本血吸虫童虫cDNA文库的免疫学筛选及阳性克隆的初步鉴定

Immunology screening and identification of Schistosoma japonicum (Chinese mainland strain)schistosomula cDNA library

目的 免疫法筛选日本血吸虫 (Schistosomajaponicum ,S j) 15d肝期童虫cDNA表达文库 ,并进行鉴定 ,以获得S j疫苗候选抗原分子。方法 采用紫外线照射致弱尾蚴免疫的兔血清 ,对S j中国大陆株 15d肝期童虫cDNA文库进行免疫学筛选 ,选取部分阳性克隆进行插入片段的核苷酸序列分析 ,将结果通过Internet送入NCBIGenBank进行同源性比较 ,并将发现的新基因送入GenBank登录。结果 对大约 10 4个噬菌斑进行了初筛 ,共获得 4 9个阳性克隆 ,复筛了其中的 12个克隆 ,获得 8个持续阳性反应克隆。对 6个克隆的核苷酸序列测定及同源性分析表明 ,2个为编码S j线粒体的基因 ,另外 4个为S j未知基因 ,新基因序列已被GenBank接受 ,并获得进入编号。 结论 从S j肝期童虫cDNA文库中筛选到一批S j基因 ,其中部分为未知基因 ,为血吸虫新的疫苗候选分子的研究提供了材料。

Aim To isolate and identify expressed Schistosoma japonicum(Chinese mainland strain)antigen genes by immunological screening schistosomula cDNA library for searching schistosome vaccine candidates Methods The cDNA library of S j schistosomula of day 15 post infection was screened with rabbit anti sera immunized by UV attennuated cercariae,and the inserts of six recombinants was sequenced Then the sequences was compared with all sequences in GenBank database by internet The novel sequences were submitted to GenBank to obtain accession numbers Results Of 10 4 phage plaques immunoscreened,49 positive cDNA clones were found,in which 12 clones were rescreened until the stock was confirmed as a pure clone Eight continuing positive clones were confirmed The sequencing data of 6 inserts showed,two was high homologous to the S j mitochondrial DNA,other four was no homologous to schistosome gene in GenBank and thus classfied as S j novel genes The sequences of the novel genes were submitted and obtained their Accession Numbers from GenBank Conclusion Immunological screening using rabbit anti sera for isolating antigen genes can been used to discover S j antigen genes with potential value for vaccine candidates