白细胞介素18受体α与β真核表达载体构建及白细胞介素18受体β稳定表达细胞株的建立  

Construction of eukaryotic expression plasmid carrying interleukin-18 receptor alpha,beta and establishment of cell line stably expressing interleukin-18 receptor beta

作　　者：陈燕[1] 赵振东[1] 

机构地区：[1]北京大学医学部基础医学院免疫系,北京市100083

出　　处：《中国组织工程研究与临床康复》2008年第28期5482-5485,共4页Journal of Clinical Rehabilitative Tissue Engineering Research

摘　　要：目的:构建人白细胞介素18受体IL-18Rα,β真核表达载体pcDNA3.1-zeocin/IL-18RαpcDNA3.0/IL-18Rβ,并将pcDNA3.0/IL-18Rβ稳定转染293细胞,构建IL-18Rβ稳定表达细胞株,用于建立IL-18信号转导的体外细胞模型。方法:实验于2006-06/2007-05在北京大学医学部免疫系实验室完成。扩增并克隆人IL-18Rα和IL-18RβcDNA,构建真核表达载体pcDNA3.1-zeocin/IL-18Rα,pcDNA3.0/IL-18Rβ。将pcDNA3.0/IL-18Rβ转染293细胞,经G418筛选建立起稳定表达IL-18Rβ的细胞株,Western-blot方法检测稳定细胞株IL-18Rβ的表达,挑选表达较高的细胞转染IL-18Rα,通过NF-κB依赖的Luciferase检测IL-18信号转导通路的建立。结果:成功构建了IL-18Rα和IL-18Rβ的真核表达载体,IL-18Rβ的基因稳定转染到了293细胞中并获得了表达,通过瞬时转染IL-18Rα真核表达载体,在293细胞中建立了IL-18信号转导的体外细胞模型。结论:成功建立起了IL-18诱导NF-κB活化的体外细胞模型,为进一步研究IL-18信号转导途径奠定了基础。AIM： To construct human interleukin-18 receptor α （IL-18R α ） and interleukin-18 receptor β （IL-18R β ） eukaryotic expression vectors named pcDNA3.1-zeocin/IL-18R α and pcDNA3.0/IL-18R β respectively, and then stably transfect pcDNA3.0/IL-18R β into 293 cells in order to establish the cell line stably expressing IL-18R β and cell model which responds IL-18 signal transduction in vitro. METHODS： The experiment was performed at the laboratory of Department of Immunology, Peking University Health Science Center from June 2006 to May 2007. DNA encoding IL-18R α or IL-18R β was amplified and cloned into pcDNA3.1-zeocin or pcDNA3.0 vector, respectively. The recombinant of pcDNA3.0/IL-18R β was transfectd into 293 cells. After screening culture by G418, stable transfected cell line was established and the expression of IL-18R β was identified by Western-blot assay. The stable cell line with the combinant eukaryotic expression vector pcDNA3.1-zeocin /IL-18Rα to establish the IL-18 signaling pathway, which was detected by NF-κ B-dependent Luciferase. RESULTS： The eukaryotic expression vectors of IL-18R α and IL-18R β were constructed successfully. The gene of IL-18R β stably expressed in 293 cells and cell model of IL-18 signaling transduction in vitro could be established in these stable 293 cell lines. CONCLUSION： The establishment of NF-κ B-activated cell model in vitro which responds to IL-18 can provide solid foundation for further experimental studies on IL-18 signaling transduction pathway.

关 键 词：IL-18Rα IL—18Rβ 293细胞 稳定细胞株 信号转导 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]