Noxa基因的克隆及其诱导A549细胞凋亡的研究  

Apoptosis of A549 Cells Induced by Cloned Noxa Gene

作　　者：陈登榜[1] 曹康[1] 杨靖[2] 潘渠[1] 陈恬[1] 李晋川[1] 王昕[1] 殷建华[1] 邹毅[3] 魏常友[1] 

机构地区：[1]成都医学院,成都610083 [2]湖北医药学院 微生物学教研室,十堰442100 [3]广东省食品药品监督管理局审评认证中心,广州510080

出　　处：《四川大学学报（医学版）》2013年第5期713-716,726,共5页Journal of Sichuan University(Medical Sciences)

基　　金：四川省教育厅面上项目(No.11ZB170);四川省卫生厅科研课题(No.120489);成都医学院校基金(No.CYZ10-004,No.CYZ11-0059);成都医学院学科建设项目(No.CYXK2012005)资助

摘　　要：目的克隆人Noxa基因,构建真核表达质粒pcDNA-Noxa,将重组质粒pcDNA-Noxa转染人肺癌A549细胞株,并观察重组质粒pcDNA-Noxa诱导A549细胞凋亡的作用。方法使用RT-PCR的方法扩增出Noxa基因,克隆至真核表达质粒pcDNA(-),构建出重组质粒pcDNA-Noxa。将重组质粒pcDNA-Noxa转染人肺癌A549细胞株,使用Western blot的方法检测Noxa基因的过表达;使用Hoechst 33258染色,观察A549细胞的凋亡;使用MTT法检测细胞活力。结果经酶切和测序鉴定,重组质粒pcDNA-Noxa构建成功。Western blot检测出Noxa基因的过表达;Hoechst 33258染色和MTT实验发现:重组质粒pcDNA-Noxa能明显诱导A549细胞出现凋亡现象。结论本实验初步探索了Noxa基因的促凋亡作用,为以后进一步深入研究Noxa基因的促凋亡功能基础。Objective To clone the Noxagene and to observe the apoptosis of A549cells transfected with the recombinant plasmid of pcDNA-Noxa.Methods The Noxa gene was obtained by PCR,and was cloned into pcDNA3.1(-).A549cells were transfected with the recombinant plasmid of pcDNA-Noxa.Western blot analysis was performed to determine the overexpression of Noxa.A549cells were stained with Hoechst 33258to observe the apoptosis.Results The recombinant plasmid of pcDNA-Noxa was successfully constructed evidenced by endonuclease digestion and sequence analysis.The overexpression of Noxa was identified using Western blot analysis.The recombinant plasmid of pcDNA-Noxa induced apoptosis of A549cells.Conclusion Noxa has exhibited potential pro-apoptotic activity against A549cells.This study is a foundation for further research into proapoptotic activity of Noxagene.

关 键 词：NOXA 克隆 真核表达 凋亡 

分 类 号：R363[医药卫生—病理学]