Thiouridine-to-Cytidine Conversion Sequencing (TUC-Seq) to Measure mRNA Transcription and Degradation Rates.
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.
['Lusser|Alexandra|A|', 'Gasser|Catherina|C|', 'Trixl|Lukas|L|', 'Piatti|Paolo|P|', 'Delazer|Isabel|I|', 'Rieder|Dietmar|D|', 'Bashin|Jeffrey|J|', 'Riml|Christian|C|', 'Amort|Thomas|T|', 'Micura|Ronald|R|']
D002460:Cell Line D003562:Cytidine D057809:HEK293 Cells D059014:High-Throughput Nucleotide Sequencing D006801:Humans D020871:RNA Stability D012333:RNA, Messenger D017423:Sequence Analysis, RNA D013194:Staining and Labeling D013891:Thiouridine D014158:Transcription, Genetic
TUC-seq RNA stability Metabolic labeling 4-Thiouridine RNA decay RNA modification Transcription rate