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Single-Molecule Analysis of Pre-mRNA Splicing with Colocalization Single-Molecule Spectroscopy (CoSMoS)

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mRNA Processing

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1648))

Abstract

Recent development of single-molecule techniques to study pre-mRNA splicing has provided insights into the dynamic nature of the spliceosome. Colocalization single-molecule spectroscopy (CoSMoS) allows following spliceosome assembly in real time at single-molecule resolution in the full complexity of cellular extracts. A detailed protocol of CoSMoS has been published previously (Anderson and Hoskins, Methods Mol Biol 1126:217–241, 2014). Here, we provide an update on the technical advances since the first CoSMoS studies including slide surface treatment, data processing, and representation. We describe various labeling strategies to generate RNA reporters with multiple dyes (or other moieties) at specific locations.

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Acknowledgments

Joerg E. Braun acknowledges funding from the Human Frontier Science Program (HFSP) LT000166/2013. Joerg E. Braun and Victor Serebrov are supported by funding to the laboratory of Melissa J. Moore: University of Massachusetts Medical School, Investigator of the Howard Hughes Medical Institute and NIH R01 GM50037.

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Authors and Affiliations

  1. RNA Therapeutics Institute, Howard Hughes Medical Institute, and Department of Biochemistry & Molecular Pharmacology, University of Massachusetts Medical School, Worcester, MA, 01605, USA

    Joerg E. Braun & Victor Serebrov

Authors
  1. Joerg E. Braun
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  2. Victor Serebrov
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Corresponding author

Correspondence to Joerg E. Braun .

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Editors and Affiliations

  1. Department of Microbiology and Molecular Genetics, School of Medicine, University of California, Irvine, California, USA

    Yongsheng Shi

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Braun, J.E., Serebrov, V. (2017). Single-Molecule Analysis of Pre-mRNA Splicing with Colocalization Single-Molecule Spectroscopy (CoSMoS). In: Shi, Y. (eds) mRNA Processing. Methods in Molecular Biology, vol 1648. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7204-3_3

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  • DOI: https://doi.org/10.1007/978-1-4939-7204-3_3

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7203-6

  • Online ISBN: 978-1-4939-7204-3

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