Probing Liquid-Liquid Phase Separation of RNA-Binding Proteins In Vitro and In Vivo

Heinrich, Stephanie and Hondele, Maria. (2022) Probing Liquid-Liquid Phase Separation of RNA-Binding Proteins In Vitro and In Vivo. In: Alternative Splicing: Methods and Protocols. New York, NY, pp. 307-333.

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Official URL: https://edoc.unibas.ch/89268/

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Biomolecular condensates and the concept of liquid-liquid phase separation (LLPS) have transformed cell biology in recent years. Condensates organize cellular content and compartmentalize biochemical reactions, in particular many processes involving RNA. This protocol is aimed at readers new to the LLPS field who want to test their protein or cellular structure of interest. We describe the basic principles of liquid-liquid phase separation, and outline initial approaches-both in vitro and in yeast cells-for the characterization of a candidate cellular condensate. First, we focus on strategies to purify phase-separating proteins and to reconstitute condensates from recombinant proteins in vitro for observation by light microscopy. Second, we describe in vivo experiments (including fluorescence recovery after photobleaching (FRAP) microscopy and 1,6-Hexanediol treatment) to test whether a subcellular structure displays liquid-like behavior in cells.
Faculties and Departments:05 Faculty of Science > Departement Biozentrum > Growth & Development > Biochemistry (Hondele)
UniBasel Contributors:Hondele, Maria
Item Type:Book Section, refereed
Book Section Subtype:Further Contribution in a Book
Series Name:Methods in Molecular Biology
Issue Number:2537
Note:Publication type according to Uni Basel Research Database: Book item
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Last Modified:23 Feb 2023 15:35
Deposited On:03 Aug 2022 10:46

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