采用荧光标记的核酶探针的微芯片电泳法检测细胞培养物中的 miRNA。
Detection of miRNA in cell cultures by using microchip electrophoresis with a fluorescence-labeled riboprobe.
机构信息
Health Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), Takamatsu, Kagawa, Japan.
出版信息
Sensors (Basel). 2012;12(6):7576-86. doi: 10.3390/s120607576. Epub 2012 Jun 7.
The analysis of a microRNA (miRNA), miR-222 isolated from the PC12 cell line, was performed by use of the ribonuclease (RNase) protection assay, cyanine 5 (Cy5)-labeled miR-222 riboprobe, and a Hitachi SV1210 microchip electrophoresis system, which can be used to evaluate the integrity of total RNA. The fluorescence intensity corresponding to the protected RNA fragment increased in a dose-dependent manner with respect to the complementary-strand RNA. More highly sensitive detection of miRNA by microchip electrophoresis than by conventional method using fluorescence-labeled riboprobe could be obtained in 180 s. An obvious increase in miR-222 expression induced by nerve growth factor in PC12 cells could be observed. These results clearly indicate the potential of microchip electrophoresis for the analysis of miRNA using RNase protection assay with a fluorescence-labeled riboprobe.
利用核糖核酸酶（RNase）保护分析、Cy5 标记的 miR-222 核糖探针以及日立 SV1210 微芯片电泳系统对从 PC12 细胞系中分离出的 microRNA（miRNA）miR-222 进行分析，该系统可用于评估总 RNA 的完整性。与互补链 RNA 相比，荧光强度与保护的 RNA 片段成正比，且随着互补链 RNA 浓度的增加而增加。与使用荧光标记的核糖探针的传统方法相比，通过微芯片电泳可以在 180 秒内实现对 miRNA 的更灵敏检测。在 PC12 细胞中，神经生长因子诱导 miR-222 表达明显增加。这些结果清楚地表明，微芯片电泳具有使用荧光标记的核糖探针进行 RNase 保护分析的 miRNA 分析的潜力。
Zotero 插件