高通量基因芯片检测阻断Cyclin B1后基因的差异表达  

Screening differentially expressed genes from Cyclin B1 down-regulated by high-throughput gene chip

作　　者：谢贤和[1] 林万尊 郑伟丽 陈婷[1] 刘俊金 

机构地区：[1]福建医科大学附属第一医院化疗科,福建福州350005

出　　处：《海南医学院学报》2018年第1期1-4,共4页Journal of Hainan Medical University

基　　金：福建省自然科学基金(2015J01457)~~

摘　　要：目的:研究Cyclin B1敲低后差异表达的基因,并筛选出与自噬相关的基因。方法:利用胸腺核苷(TdR)双阻断法使鼻咽癌细胞CNE-2周期同步化至S期,siRNA干扰Cyclin B1的表达,流式细胞术验证转染效率,q-PCR和western blot验证siRNA沉默效率,高通量基因芯片筛选对照组和实验组差异表达的基因。结果:经2.5mmol/L的TdR同步化后,流式细胞仪检测细胞周期,获得S期细胞。用脂质体转染法将FAM-siRNA导入CNE-2细胞后,流式细胞术检测转染效率为85.6%,将Cyclin B1-siRNA导入CNE-2细胞后,Cyclin B1的mRNA和蛋白表达均较对照组明显下降85%(P<0.01)。基因芯片结果显示与对照组相比,转染Cyclin B1-siRNA后一共有2 408个差异表达的基因,其中1 245个基因显著上调,1 163个基因显著下调,初步筛选出上调的自噬相关基因PTEN。结论:Cyclin B1-siRNA可以有效地沉默Cyclin B1蛋白的表达,并可以使2408个基因差异表达,初步筛选出上调的自噬相关基因PTEN。Objective:The study aimed to screen differentially expressed genes by silencing Cyclin B1,and to sift out autophagy-related genes.Methods:Double thymidine deoxyribonucleoside(TdR)blcoking was used to synchronize nasopharyngeal carcinoma cell(CNE-2)to S phase,then flow cytometry was applied to test transfection efficiency.The mRNA and protein expression level of Cyclin B1 was assessed by q-PCR and western blot,respectively.Differentially expressed genes were screened by high-throughput gene chip.Results:Double TdR(2.5 mmol/L)blocking was used to synchronize the cell cycle to S phase.The transfection efficiency of CNE-2 cells was 87%.Compared with negative group,Cyclin B1-siRNA treated group significantly down-regulated mRNA expression of Cyclin B1(80%)and protein level(75.3%)(P<0.01).Totally,2408 differentially expressed genes were found in CNE-2,including 1245 up-regulated genes and 1163 down-regulated genes.Moreover,PTEN,an autophagy-related gene,was preliminarily sifted out.Conclusions:Cyclin B1-siRNA significantly down-regulated the expression of Cyclin B1 and yielded a total of 2408 differentially expressed genes,including PETN(an autophagy-related gene).

关 键 词：CYCLIN B1 细胞周期同步化 高通量基因芯片 自噬相关基因 

分 类 号：R730[医药卫生—肿瘤] R34[医药卫生—临床医学]