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Methods to Study Repeat Fragility and Instability in Saccharomyces cerevisiae

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Genome Instability

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1672))

Abstract

Trinucleotide repeats are common in the human genome and can undergo changes in repeat length. Expanded CAG repeats have been linked to over 14 human diseases and are considered hotspots for breakage and genomic rearrangement. Here, we describe two Saccharomyces cerevisiae based assays that evaluate the rate of chromosome breakage that occurs within a repeat tract (fragility), and a PCR-based assay to evaluate tract length changes (instability). The first fragility assay utilizes end-loss and subsequent telomere addition as the main mode of repair of a yeast artificial chromosome (YAC). The second fragility assay relies on the fact that a chromosomal break stimulates recombination-mediated repair. In addition to understanding the role of fragility at repetitive DNA sequences, both assays can be modified to evaluate instability of a CAG repeat using a PCR-based assay. All three assays have been essential in understanding the genetic mechanisms that cause chromosome breaks and tract-length changes at unstable repeats.

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Acknowledgments

Work in our laboratory is currently supported by grants from the National Institute of Health (GM105473) and National Science Foundation (MCB 1330743). We would like to thank past and present members of the C. H. Freudenreich and V.A. Zakian labs for their contributions to the development of these assays.

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Authors and Affiliations

  1. Department of Biology, Tufts University, 200 Boston Avenue, Medford, MA, 02155, USA

    Erica J. Polleys & Catherine H. Freudenreich

Authors
  1. Erica J. Polleys
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  2. Catherine H. Freudenreich
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Corresponding author

Correspondence to Catherine H. Freudenreich .

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Editors and Affiliations

  1. Dipartimento di Bioscienze, Università degli Studi di Milano, Milano, Milano, Italy

    Marco Muzi-Falconi

  2. Donnelly Centre and Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada

    Grant W Brown

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Polleys, E.J., Freudenreich, C.H. (2018). Methods to Study Repeat Fragility and Instability in Saccharomyces cerevisiae . In: Muzi-Falconi, M., Brown, G. (eds) Genome Instability. Methods in Molecular Biology, vol 1672. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7306-4_28

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  • DOI: https://doi.org/10.1007/978-1-4939-7306-4_28

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7305-7

  • Online ISBN: 978-1-4939-7306-4

  • eBook Packages: Springer Protocols

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