A translational repression reporter assay for the analysis of RNA-binding protein consensus sites
| Autor: | Jessica Nowacki, Mateo Malenica, Stefan Schmeing, Damian Schiller, Benjamin Buchmuller, Gulshan Amrahova, Peter ‘t Hart |
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| Jazyk: | angličtina |
| Rok vydání: | 2023 |
| Předmět: | |
| Zdroj: | RNA Biology, Vol 20, Iss 1, Pp 85-94 (2023) |
| Druh dokumentu: | article |
| ISSN: | 1547-6286 1555-8584 15476286 |
| DOI: | 10.1080/15476286.2023.2192553 |
| Popis: | RNA-binding proteins are essential regulators of RNA processing and function. Translational repression assays can be used to study how they interact with specific RNA sequences by insertion of such a consensus sequence into the 5’ untranslated region of a reporter mRNA and measuring reporter protein translation. The straightforward set-up of these translational repression assays avoids the need for the isolation of the protein or the RNA providing speed, robustness and a low-cost method. Here, we report the optimization of the assay to function with linear RNA sequences instead of the previously reported hairpin type sequences to allow the study of a wider variety of RNA-binding proteins. Multiplication of a consensus sequence strongly improves the signal allowing analysis by both fluorescence intensity measurements and flow cytometry. |
| Databáze: | Directory of Open Access Journals |
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