5'-溴尿苷IP追踪（BRIC）-Seq法测定RNA半衰期

5'-Bromouridine IP Chase (BRIC)-Seq to Determine RNA Half-Lives.

作者信息

Yamada Toshimichi, Imamachi Naoto, Onoguchi-Mizutani Rena, Imamura Katsutoshi, Suzuki Yutaka, Akimitsu Nobuyoshi

机构信息

Isotope Science Center, The University of Tokyo, Tokyo, Japan.

Department of Microbiology and Molecular Genetics, Graduate School of Pharmaceutical Sciences, Chiba University, Chiba, Japan.

出版信息

Methods Mol Biol. 2018;1720:1-13. doi: 10.1007/978-1-4939-7540-2_1.

DOI:10.1007/978-1-4939-7540-2_1

PMID:29236247

Abstract

Analysis of RNA stability at genome-wide level is an advanced method in RNA biology that examines the half-life of each transcript. In particular, a pulse-labeling method using uridine analogs enables the determination of half-life of each transcript under physiologically undisturbed conditions. The technique involves pulse labeling of endogenous RNAs in mammalian cells with 5'-bromouridine (BrU), followed by measuring the chronological decrease of BrU-labeled RNAs using deep sequencing (BRIC-seq). Here, we describe a detailed protocol and technical tips for BRIC-seq.

摘要

全基因组水平的RNA稳定性分析是RNA生物学中的一种先进方法，用于检测每个转录本的半衰期。特别是，使用尿苷类似物的脉冲标记方法能够在生理未受干扰的条件下测定每个转录本的半衰期。该技术包括用5'-溴尿苷（BrU）对哺乳动物细胞中的内源性RNA进行脉冲标记，然后使用深度测序（BRIC-seq）测量BrU标记RNA随时间的减少情况。在此，我们描述了BRIC-seq的详细方案和技术要点。