胞苷
抄写(语言学)
核糖核酸
信使核糖核酸
生物
核苷
计算生物学
细胞生物学
基因
化学
生物化学
语言学
哲学
酶
作者
Alexandra Lusser,Catherina Gasser,Lukas Trixl,Paolo Piatti,Isabel Delazer,Dietmar Rieder,Jeffrey Bashin,Christian Riml,Thomas Amort,Ronald Micura
出处
期刊:Methods in molecular biology
日期:2019-11-26
卷期号:: 191-211
被引量:19
标识
DOI:10.1007/978-1-4939-9822-7_10
摘要
The study of RNA dynamics, specifically RNA transcription and decay rates, has gained increasing attention in recent years because various mechanisms have been discovered that affect mRNA half-life, thereby ultimately controlling protein output. Therefore, there is a need for methods enabling minimally invasive, simple and high-throughput determination of RNA stability that can be applied to determine RNA transcription and decay rates in cells and organisms. We have recently developed a protocol which we named TUC-seq to directly distinguish newly synthesized transcripts from the preexisting pool of transcripts by metabolic labeling of nascent RNAs with 4-thiouridine (4sU) followed by osmium tetroxide-mediated conversion of 4sU to cytidine (C) and direct sequencing. In contrast to other related methods (SLAM-seq, TimeLapse-seq), TUC-seq converts 4sU to a native C instead of an alkylated or otherwise modified nucleoside derivative. TUC-seq can be applied to any cell type that is amenable to 4sU labeling. By employing different labeling strategies (pulse or pulse-chase labeling), it is suitable for a broad field of applications and provides a fast and highly efficient means to determine mRNA transcription and decay rates.
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