|
|||||
|
|
| 杜氏利什曼原虫四川分离株无鞭毛体蛋白基因的克隆及真核表达 | |
| 引用本文： | 李金福,陈建平,田玉,杨志伟,马莹,胡孝素. 杜氏利什曼原虫四川分离株无鞭毛体蛋白基因的克隆及真核表达[J]. 中国寄生虫学与寄生虫病杂志, 2007, 25(2): 124-128 |
| 作者姓名： | 李金福 陈建平 田玉 杨志伟 马莹 胡孝素 |
| 作者单位： | 四川大学华西基础医学与法医学院寄生虫学教研室,成都610041 |
| 基金项目： | 国家自然科学基金资助（No.39870656） |
| 摘 要： | 目的 克隆、真核表达杜氏利什曼原虫四川株的无鞭毛体蛋白（amastin）编码基因。 方法 PCR扩增杜氏利什曼原虫四川汶川人株L.d.SC10H2与四川南坪犬株L.d.SC7的无鞭毛体蛋白基因，将该基因导入pcDNA3.1（+），构建真核表达重组质粒，转染NIH3T3细胞，采用免疫荧光法鉴定重组质粒的瞬时表达；RT-PCR和Western blotting鉴定稳定表达。 结果 2株杜氏利什曼原虫均扩增出552 bp的无鞭毛体蛋白基因，同源性为86％。转染后在NIH3T3细胞膜和细胞内观察到较强的绿色荧光，表明无鞭毛体蛋白基因在NIH3T3细胞中获得短暂表达。细胞裂解产物经Western blotting，在相对分子质量（Mr）约20 000处检测到阳性杂交信号，表明无鞭毛体蛋白基因在NIH3T3细胞内获得了稳定表达。 结论 获得了我国杜氏利什曼原虫四川分离株L.d.SC10H2和L.d.SC7的无鞭毛体蛋白基因序列，并在NIH3T3细胞中稳定表达。 |
| 关 键 词： | 杜氏利什曼原虫 无鞭毛体蛋白 基因克隆 表达 |
| 文章编号： | 1000-7423（2007）-02-0124-05 |
| 收稿时间： | 2006-09-04 |
| 修稿时间： | 2006-09-04 |
Cloning of Amastin Gene of Leishmania donovani Isolates from Sichuan Province and its Expression in Eucaryotic System |
|
| LI Jin-fu,CHEN Jian-ping,TIAN Yu,YANG Zhi-wei,MA Ying,HU Xiao-su. Cloning of Amastin Gene of Leishmania donovani Isolates from Sichuan Province and its Expression in Eucaryotic System[J]. Chinese Journal of Parasitology and Parasitic Diseases, 2007, 25(2): 124-128 | |
| Authors: | LI Jin-fu CHEN Jian-ping TIAN Yu YANG Zhi-wei MA Ying HU Xiao-su |
| Affiliation: | Department of Parasitology, School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China |
| Abstract: | OBJECTIVE: To clone and express the amastin gene of two Leishmania donovani isolates from Sichuan Province of China. METHODS: Amastin gene was amplified from nuclear DNA of two L.donovani isolates, cloned into pcDNA3.1(+), and sequenced by the dideoxy chain termination. NIH3T3 cell was transfected by recombinant plasmid. Transient expression of amastin gene was detected by immunofluoresence and stable expression was detected by RT-PCR and Western blot. RESULTS: Amastin gene of both isolates was 552 bp. Sequence analysis showed that the similarity was 86% between the two isolates. A high green fluorescence was found on the cell membrane and inside the cell. The NIH3T3 cell was transfected by the recombinant plasmid successfully. Amastin gene was obtained by RT-PCR from the transfected NIH3T3 cells. Western blot analysis showed that there was a protein about Mr 20 000 in lysate of the transfected NIH3T3 cells, indicating that the amastin gene was expressed in the cells. CONCLUSION: Amastin gene of the two L.donovani isolates has been cloned and the gene can be expressed stably in the NIH3T3 cell. |
| Keywords: | Leishmania donovani Amastin Gene cloning Expression |
| 本文献已被 维普 等数据库收录！ | |
| 点击此处可从《中国寄生虫学与寄生虫病杂志》浏览原始摘要信息 | |
| 点击此处可从《中国寄生虫学与寄生虫病杂志》下载全文 | |