Construction of Eukaryotic Expression Vector with Partial Encoding Sequence of Actin from Cryptosporidium andersoni  

安氏隐孢子虫肌动蛋白部分编码基因真核表达载体的构建(英文)

作　　者：陈健[1,2] 胡进平[1,2] 宫鹏涛[2] 李建华[2] 杨举[2] 李赫[2] 张国才[2] 张西臣[2] 任文陟[1] 

机构地区：[1]吉林大学实验动物中心,吉林长春130062 [2]吉林大学畜牧兽医学院,吉林长春130062

出　　处：《Agricultural Science & Technology》2012年第3期641-643,655,共4页农业科学与技术（英文版）

基　　金：Supported by the National Key Technology R & D Program (2007BAD40B05),China;the National High Technology Research and Development Program of China(2006AA10A207)~~

摘　　要：[Objective] To clone the actin gene of Cryptosporidium andersoni, and to study its eukaryotic expression in Hela cells. [Methed] Specific primers were designed for the partial encoding sequence of actin, which were obtained by screening the T7 phage display library of Cryptosporidium andersoni, and the actin gene CA42 was amplified by PCR. Recombinant eukaryotic expression plasmid pVAX1-CA42 was constructed and transfected to Hela cells with lipofection strategy. Indirect im- munofluorescence staining, SDS-PAGE and Western blotting analysis were used to detect the expression of recombinant protein in Hela cells. [Result] CA42 protein was successfully expressed in Hela cells, and the expression products had reactogenicity. [Conclusion] The partial encoding sequence of actin from Cryptosporidium andersoni has been successfully cloned, and it can be stably expressed in Hela Cells[目的]将安氏隐孢子虫肌动蛋白基因进行克隆和在Hela细胞中真核表达。[方法]根据筛选安氏隐孢子虫T7噬菌体展示文库获得的肌动蛋白(CA42)部分编码基因序列设计特异性引物,扩增目的基因CA42,构建重组真核表达载体pVAX1-CA42,将其转化入Hela细胞,用间接免疫荧光、SDS-PAGE、Western blotting检测CA42蛋白在转染细胞中的表达情况。[结果]重组质粒能在Hela细胞中表达,且表达产物具有反应原性。[结论]成功克隆并表达了安氏隐孢子虫肌动蛋白基因,并在Hela细胞中能够稳定表达。

关 键 词：Cryptosporidium anderssonr ACTIN Eukaryotic expression 

分 类 号：S852.7[农业科学—基础兽医学]