摘要
目的:克隆人心房肌小电导钙激活钾通道亚型2(SK2),构建稳定表达SK通道的HEK293细胞系并研究其基本电生理学特性。方法:以人心房肌为标本,通过逆转录,重叠PCR法和定向克隆构建真核表达载体pIRES-hrGFP-SK2,采用脂质体法转染至HEK293细胞并筛选稳定表达的细胞系;使用全细胞和单通道膜片钳技术进行电流记录,研究其基本电生理学特性。结果:全细胞和单通道膜片钳均能在表达的HEK293细胞上记录到SK2通道电流,其电流具有明显的胞内钙离子依赖性。结论:成功构建稳定表达人心房肌SK2通道的HEK293细胞系,为以后研究通道功能活动奠定了基础。
Objective： To clone the small conductance calcium activated potassium channels type 2（SK2）of human atrial tissue,construct stable HEK293 cells expressed SK2 and investigate the basic electrophysiological properties.Methods： The expression plasmid of pIRES-hrGFP-SK2 was constructed by reverse transcription,overlapping PCR and site-directed cloning.The plasmid was transfected by Lipofectamine 2000 and stable cell lines expressed SK2 were selected.The basic properties of SK2 were investigated by using patch clamp technique.Results： SK2 currents could be recorded in HEK293 cells expressed SK2 channels in whole-cell and single channel configuration.SK2 currents were calcium dependenent.Conclusion： Stable HEK293 cells expressed SK2 are constructed successfully,which establishes the basis for further research about the regulation of SK2 channels.
出处
《泸州医学院学报》
2013年第3期203-206,共4页
Journal of Luzhou Medical College
基金
国家自然科学基金项目(30870903)
四川省科技计划(2011FZ0106)
关键词
小电导钙激活钾通道
基因转染
心房颤动
Small conductance calcium activated potassium channels
Gene transfection
Atrial fibrillation
