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Analysis of RNA Folding and Ribonucleoprotein Assembly by Single-Molecule Fluorescence Spectroscopy

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Spectroscopic Methods of Analysis

Part of the book series: Methods in Molecular Biology ((MIMB,volume 875))

Abstract

To execute their diverse range of biological functions, RNA molecules must fold into specific tertiary structures and/or associate with one or more proteins to form ribonucleoprotein (RNP) complexes. Single-molecule fluorescence spectroscopy is a powerful tool for the study of RNA folding and RNP assembly processes, directly revealing different conformational subpopulations that are hidden in conventional ensemble measurements. Moreover, kinetic processes can be observed without the need to synchronize a population of molecules. In this chapter, we describe the fluorescence spectroscopic methods used for single-molecule measurements of freely diffusing or immobilized RNA molecules or RNA–protein complexes. We also provide practical protocols to prepare the fluorescently labeled RNA and protein molecules required for such studies. Finally, we provide two examples of how these various preparative and spectroscopic methods are employed in the study of RNA folding and RNP assembly processes.

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Acknowledgments

Work in the author’s laboratory was supported by NIH grants RO1 GM058873 and P50 GM082545.

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Authors and Affiliations

  1. Department of Molecular Biology, The Scripps Research Institute, La Jolla, CA, USA

    Goran Pljevaljčić, Rae Robertson-Anderson, Edwin van der Schans & David Millar

Authors
  1. Goran Pljevaljčić
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  2. Rae Robertson-Anderson
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  3. Edwin van der Schans
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  4. David Millar
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Corresponding author

Correspondence to David Millar .

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Editors and Affiliations

  1. Dept. Biochemistry & Molecular Biology, University of Texas Medical Branch, University Blvd. 301, Galveston, 77555-1053, Texas, USA

    Wlodek M. Bujalowski

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Pljevaljčić, G., Robertson-Anderson, R., van der Schans, E., Millar, D. (2012). Analysis of RNA Folding and Ribonucleoprotein Assembly by Single-Molecule Fluorescence Spectroscopy. In: Bujalowski, W. (eds) Spectroscopic Methods of Analysis. Methods in Molecular Biology, vol 875. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-806-1_15

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  • DOI: https://doi.org/10.1007/978-1-61779-806-1_15

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-61779-805-4

  • Online ISBN: 978-1-61779-806-1

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