Huang Jinliang, Wang Geng
Ministry of Education Key Laboratory of Bioinformatics, Cell Biology and Development Center, School of Life Sciences, Tsinghua University, Beijing 100084, China.
Bio Protoc. 2019 Jun 5;9(11):e3247. doi: 10.21769/BioProtoc.3247.
Mitochondria have two sets of RNAs. One is encoded in mitochondrial genome, and the other that consists of imported RNAs within mitochondria and cytosolic RNAs associated with mitochondrial outer membrane is encoded in the nucleus. These mitochondrial RNAs play important roles in mitochondrion biosynthesis and signaling in and out of mitochondria. Isolation and analysis of mitochondrial RNAs can provide useful information on understanding the mitochondrial regulation of cellular processes. However, several ribonuclease activities have been found in mitochondria, which will degrade mitochondrial RNAs during the isolation process if they are not properly inactivated. Here, we describe an improved method to inactivate the ribonuclease activities prior to RNA extraction, and thus provide a reliable protocol to isolate mammalian mitochondrial RNAs for quantitative RT-PCR and other assays.
线粒体有两套RNA。一套由线粒体基因组编码，另一套由线粒体内导入的RNA和与线粒体外膜相关的胞质RNA组成，由细胞核编码。这些线粒体RNA在线粒体生物合成以及线粒体内外的信号传导中发挥重要作用。线粒体RNA的分离和分析可为理解细胞过程的线粒体调控提供有用信息。然而，在线粒体中发现了几种核糖核酸酶活性，如果在分离过程中未适当使其失活，它们会降解线粒体RNA。在此，我们描述了一种在RNA提取之前使核糖核酸酶活性失活的改进方法，从而提供了一种可靠的方案来分离用于定量RT-PCR和其他检测的哺乳动物线粒体RNA。
