Meiosis pp 117-142

Part of the Methods in Molecular Biology book series (MIMB, volume 557)

Gel Electrophoresis Assays for Analyzing DNA Double-Strand Breaks in Saccharomyces cerevisiae at Various Spatial Resolutions

  • Hajime Murakami
  • Valérie Borde
  • Alain Nicolas
  • Scott Keeney
Protocol

Abstract

Meiotic recombination is triggered by programmed DNA double-strand breaks (DSBs), which are catalyzed by Spo11 protein in a type II topoisomerase-like manner. Meiotic DSBs can be detected directly using physical assays (gel electrophoresis, Southern blotting, and indirect end-labeling) applied to samples of genomic DNA from sporulating cultures of budding and fission yeast. Such assays are extremely useful for quantifying and characterizing many aspects of the initiation of meiotic recombination, including the timing of DSB formation relative to other events, the distribution of DSBs across the genome, and the influence on DSB formation of mutations in recombination factors and other gene products. By varying the type of gel electrophoresis and other parameters, the spatial resolution of DSB analysis can range from single nucleotides up to whole yeast chromosomes.

Key words

yeast meiotic recombination Southern blotting pulsed-field gel electrophoresis 

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Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2009

Authors and Affiliations

  • Hajime Murakami
    • 1
  • Valérie Borde
    • 2
  • Alain Nicolas
    • 3
  • Scott Keeney
    • 4
  1. 1.Molecular Biology ProgramMemorial Sloan-Kettering Cancer CenterNew YorkUSA
  2. 2.Institut Curie, Centre de Recherche, UMR7147-CNRS, UniversitéPierre et Marie CurieFrance
  3. 3.Institut Curie, Centre de Recherche, UMR7147-CNRS, UniversitéPierre et Marie CurieFrance
  4. 4.Howard Hughes Medical Institute and Molecular Biology ProgramMemorial Sloan-Kettering Cancer CenterNew YorkUSA

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