Dynamic Evolution of Poly-A Tail Lengths Visualized by RNAse H Assay and Northern Blot Using Nonradioactive Probes in Yeast.
| Autor: | Audebert L; Department of Microbiology and Molecular Medicine, Faculty of Medicine, University of Geneva, Institute of Genetics and Genomics of Geneva, Geneva, Switzerland. Lena.Audebert@unige.ch.; Institut Pasteur, Université Paris Cité, CNRS UMR3525, Genetics of Macromolecular Interactions, Paris, France. Lena.Audebert@unige.ch.; Sorbonne Université, Collège doctoral, Paris, France. Lena.Audebert@unige.ch., Saveanu C; Institut Pasteur, Université Paris Cité, RNA Biology of Fungal Pathogens, Paris, France.; Institut Pasteur, Université Paris Cité, CNRS UMR3525, Genetics of Macromolecular Interactions, Paris, France., Collart MA; Department of Microbiology and Molecular Medicine, Faculty of Medicine, University of Geneva, Institute of Genetics and Genomics of Geneva, Geneva, Switzerland. |
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| Jazyk: | angličtina |
| Zdroj: | Methods in molecular biology (Clifton, N.J.) [Methods Mol Biol] 2025; Vol. 2863, pp. 45-60. |
| DOI: | 10.1007/978-1-0716-4176-7_4 |
| Abstrakt: | Poly-A tail length dynamics have been extensively studied from yeast to human, mostly using reporter transcripts. Recent studies have been carried out genome-wide to determine the status of poly-A tails at steady state. However, poly-A tail measurement at equilibrium gives an overall length that reflects a mixture of the different poly-A tail sizes for a single transcript. New genome-scale techniques are emerging to estimate dynamic of poly-A tails lengths, but they are not yet routine and individual validation experiments are useful. In this chapter we describe a protocol for visualizing poly-A tail lengths following transcription inhibition for a reporter mRNA using denaturing poly-acrylamide gel electrophoresis and northern blot assay. This protocol is quick to set up, requires the purchase of only a few specific reagents, does not rely on radioactivity for RNA monitoring, and can be easily implemented in any molecular biology laboratory. (© 2025. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.) |
| Databáze: | MEDLINE |
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