日本血吸虫大陆株卵黄铁蛋白基因重组及其真核表达  

SCHISTOSOMA JAPONICUM CHINESE STRAIN: CLONING AND EUKARYOTIC EXPRESSION OF THE YOLK FERRITIN GENE

作　　者：周俊梅[1] 余新炳[1] 吴忠道[1] 李焱[1] 郑亦男[1] 

机构地区：[1]中山医科大学寄生虫学教研室,广东广州510089

出　　处：《中国寄生虫病防治杂志》2001年第3期204-206,共3页Chinese Journal of Parasitic Disease Control

基　　金：中山医科大学"211工程"重点学科建设基金资助 (No.981 69)

摘　　要：目的　构建日本血吸虫卵黄铁蛋白基因 (Sj Yol Fer)的真核表达重组质粒 p L Yol Fer,转染 Hela细胞进行表达 ,并对表达产物进行鉴定。　方法　用 PCR法从日本血吸虫大陆株成虫 c DNA文库中扩增目的基因片段 ,经 RNA斑点杂交显示转录差异 ,然后定向克隆入表达载体 p L XSN。转染重组体到 Hela细胞中进行表达 ,并用 SDS- PAGE和 Westernblot分析表达产物。　结果　PCR扩增产物大小约 6 0 0 bp左右 ,与雌虫 RNA的杂交信号明显强于雄虫。获得重组质粒p L Yol Fer,特异性表达产物约为 2 1ku,能被日本血吸虫感染兔血清及小鼠血清识别。　结论　为进一步免疫原性实验奠定了基础。Objective To construct the eukaryotic expressed recombinant plasmid pLYolFer which contains Chinese Schistosoma japonicum Yolk ferritin cDNA, exam its transcript by RNA dot blot, express it in Hela cells and identify its expression condition. Methods The target cDNA fragment was amplified from the adult S. japonicum Chinese strain cDNA library by PCR. RNA dot blot was done and the fragment was ligated into the EcoRⅠ and BamHⅠ site in the vector pLXSN by double restrictive enzymes reaction and linking reaction. The recombinants was transfected into Hela cells using Lipofectamine. The expressed products were examined by SDS-PAGE and Western blot. Results The cDNA fragment was about 600 bp. The recombinant plasmid was constructed successfully. The special expressing product was 21 ku which could be recognized by antibodies in sera of rabbit or mouse infected with S. japonicum. Conclusion The eukaryotic expressed recombinant plasmid pLYolFer which contains Chinese S. japonicum Yolk ferritin cDNA was constructed. It can be used in DNA vaccination further.

关 键 词：卵黄铁蛋白 日本血吸虫 基因克隆 基因表达 

分 类 号：R383.24[医药卫生—医学寄生虫学]