Ebhardt H Alexander, Ovando Mariana Oviedo, Unrau Peter J
Institute of Molecular Systems Biology, Eidgenössische Technische Hochschule Zürich, Zürich, Switzerland.
Methods Mol Biol. 2012;894:223-39. doi: 10.1007/978-1-61779-882-5_15.
Small RNAs, defined as noncoding 20-30-nt-long RNAs, are instrumental regulators of cellular processes in most eukaryotes. In this chapter we describe three methods for extracting small RNA from cells: a general method, one plant specific and a third particular to conifers. Further, protocols are given for the analysis and quantification of small RNAs using polyacrylamide gel-based approaches. A native streptavidin gel-shift assay, useful for measuring the relative amounts of multiple small RNAs simultaneously, is presented. To further characterize small RNAs biochemically, a sodium periodate assay probing for 2', 3' hydroxyl groups on the 3' terminus of small RNAs is outlined.
小RNA被定义为长度为20 - 30个核苷酸的非编码RNA，在大多数真核生物中是细胞过程的重要调节因子。在本章中，我们描述了三种从细胞中提取小RNA的方法：一种通用方法、一种植物特异性方法和第三种针叶树特有的方法。此外，还给出了使用基于聚丙烯酰胺凝胶的方法分析和定量小RNA的方案。介绍了一种天然链霉亲和素凝胶迁移试验，该试验可用于同时测量多种小RNA的相对含量。为了进一步从生化角度表征小RNA，概述了一种用于探测小RNA 3'末端2'，3'羟基的高碘酸钠试验。
