应用锁式探针滚环扩增检测小鼠畸胎瘤分化过程中单细胞基因表达水平  

In situ analysis of gene expression in single cell with Padlock-RCA

作　　者：杜洪震[1] 常德[1] 李利民[1] 

机构地区：[1]中国医学科学院基础医学研究所北京协和医学院基础学院病理系,北京100005

出　　处：《基础医学与临床》2011年第5期556-560,共5页Basic and Clinical Medicine

摘　　要：目的应用锁式探针介导的滚环扩增技术(Padlock-RCA)原位检测干细胞分化过程中单细胞mRNA表达水平。方法利用维甲酸(RA)诱导小鼠畸胎瘤细胞F9分化模型,应用锁式探针滚环扩增原位基因检测技术检测Nanog在其分化前后单细胞mRNA表达水平,结合实时定量RT-PCR分析该方法的效果。结果锁式探针滚环扩增原位基因表达检测方法能够检测单个细胞mRNA表达水平,并能够有效的反映F9细胞分化前较分化后Nanog基因表达水平显著降低(P<0.05)。结论成功应用锁式探针滚环扩增原位基因表达检测技术分析Nanog基因在F9分化前后单细胞中表达水平,为进一步研究单细胞基因表达及细胞亚群功能分析提供了一种可靠的新方法。Objective To apply Padlock rolling-circle amplification（Padlock-RCA） for the analysis of in situ gene expression in individual cells during differentiation.Methods Retinoic acid induced differentiation of mouse teratocarcinoma F9 model was used,Nanog mRNA expression in F9 cells before and after the induced differentiation was analyzed with Padlock-RCA and compared with result of RT-PCR.Results Padlock-RCA can efficiently detect the level of Nanog mRNA expression in single cells.Nanog specific Padlock-RCA products were significantly decreased in F9 cell following retinoic acid induction,and confirmed with RT-PCR（P0.05）.Conclusion We have successfully applied Padlock-RCA to analyze differential Nanog mRNA expressions before and after the induced differentiation in single cell,and provided a reliable method for molecular and functional analyses of gene expression in single cell or cell subpopulations.

关 键 词：锁式探针滚环扩增 小鼠畸胎瘤细胞 MRNA 分化 

分 类 号：R730.2[医药卫生—肿瘤]