中国寄生虫学与寄生虫病杂志 ›› 2000, Vol. 18 ›› Issue (6): 7-346.

• 论著 • 上一篇    下一篇

恶性疟原虫己糖转运体编码基因的体外扩增、克隆及表达

魏泉德 1;余新炳2;叶苓2;徐劲2
  

  1. 广东医学院寄生虫学教研室,湛江 524023 中山医科大学寄生虫学教研室,广州 510089
  • 收稿日期:1900-01-01 修回日期:1900-01-01 出版日期:2000-12-30 发布日期:2000-12-30

AMPLIFICATION, CLONING AND EXPRESSION OF A GENE ENCODING HEXOSE TRANSPORTER OF PLASMODIUM FALCIPARUM

WEI Quan-de 1;YU Xin-bing 2;YE Ling 2;XU Jin 2
  

  1. 1 Department of Parasitology;Guangdong Medical College;Zhanjiang 524023; 2 Department of Parasitology;Sun Yat-sen University of Medical Sciences;Guangzhou 510089
  • Received:1900-01-01 Revised:1900-01-01 Online:2000-12-30 Published:2000-12-30

摘要:   [目的 ]体外扩增、克隆和表达恶性疟原虫海南分离株的己糖转运体 (PfHT1)基因 ,为研究其保护性免疫创造条件。 [方法 ]恶性疟原虫FCC1/HN株的体外培养 ;碱裂解法提取基因组DNA ;PfHT1基因的PCR扩增、克隆 ;脂质体介导法转染HEPG2细胞株及真核表达。 [结果 ]从恶性疟原虫海南分离株基因组DNA中扩增出特异性的编码PfHT1的基因序列 ,片段大小为 15 16bp ;成功构建 pN3 HT1真核表达重组质粒并在肝癌细胞HEPG2中稳定表达。 [结论 ]体外成功扩增、克隆恶性疟原虫PfHT1编码序列 ;重组质粒转染成功并获稳定表达融合蛋白的 pN3 HT1/HEPG2细胞株

关键词: 恶性疟原虫, 己糖转运体, 己糖转运体基因, 克隆, 表达

Abstract:  Objective] To amplify,clone and express of a gene encoding hexose transporter of Plasmodium falcipuram(PfHT1) from Southern China isolate FCC1/HN for studing the immune of recombinant which protective from malaria parasite infection. [Methods] Cultivation of P.falciparum isolate FCC1/HN in vitro; extraction of genomic DNA from FCC1/HN using the alkali specific cleavage method; PCR amplification of PfHT1 and cloning into eukaryotic expression vector, pEGFPN3. The recombinant was introduced into mammalian cells, HEPG2 by using liposome-mediated transfection. [Results] The gene encoding PfHT1 was specifically amplified from the genomic DNA of P.falciparum isolate FCC1/HN. The size of amplified fragment was 1 516 base pair. The eukaryotic expression recombinant, pN3-HT1 , was constructed and expressed steadily in the hepatocarcinoma cell lines, HEPG2. [Conclusion] The gene encoding PfHT1 was successfully amplified and cloned. The pN3-HT1/HEPG2 cell line was built for expressing fusion protein of GFP-HT1.

Key words: Plasmodium falciparum, hexose transporter, gene of PfHT1, clone, expression