Data Analysis Pipeline for Detection and Quantification of Pseudouridine (ψ) in RNA by HydraPsiSeq.
| Autor: | Pichot F; Institute of Pharmacy of Pharmacy and Biochemistry, Johannes Gutenberg University Mainz, Mainz, Germany.; Université de Lorraine, CNRS, INSERM, UAR2008/US40 IBSLor, EpiRNA-Seq Core facility, F-54000, Nancy, France., Marchand V; Université de Lorraine, CNRS, INSERM, UAR2008/US40 IBSLor, EpiRNA-Seq Core facility, F-54000, Nancy, France., Helm M; Institute of Pharmacy of Pharmacy and Biochemistry, Johannes Gutenberg University Mainz, Mainz, Germany., Motorin Y; Université de Lorraine, CNRS, INSERM, UAR2008/US40 IBSLor, EpiRNA-Seq Core facility, F-54000, Nancy, France. Yuri.Motorin@univ-lorraine.fr.; Université de Lorraine, CNRS, UMR7365 IMoPA, F-54000, Nancy, France. Yuri.Motorin@univ-lorraine.fr. |
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| Jazyk: | angličtina |
| Zdroj: | Methods in molecular biology (Clifton, N.J.) [Methods Mol Biol] 2023; Vol. 2624, pp. 207-223. |
| DOI: | 10.1007/978-1-0716-2962-8_14 |
| Abstrakt: | Pseudouridine, a modified RNA residue formed by the isomerization of its parental U nucleotide, is prevalent in a majority of cellular RNAs; its presence was reported in tRNA, rRNA, and sn/snoRNA as well as in mRNA/lncRNA. Multiple analytical deep sequencing-based approaches have been proposed for pseudouridine detection and quantification, among which the most popular relies on the use of soluble carbodiimide (termed CMCT). Recently, we developed an alternative protocol for pseudouridine mapping and quantification. The principle is based on protection of pseudouridine against random RNA cleavage by hydrazine/aniline treatment (HydraPsiSeq protocol). This "negative" detection mode requires higher sequencing depth and provides a precise quantification of the pseudouridine content. All "wet-lab" technical details of the HydraPsiSeq protocol have been described in recent publications. Here, we describe all bioinformatics analysis steps required for data processing from raw reads to the pseudouridylation profile of known or unknown RNA. (© 2023. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.) |
| Databáze: | MEDLINE |
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