Meiosis pp 197-207

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

Detection of SPO11-Oligonucleotide Complexes from Mouse Testes

  • Jing Pan
  • Scott Keeney
Protocol

Abstract

The SPO11 protein generates programmed DNA double-strand breaks (DSBs) that initiate meiotic recombination. Endonucleolytic cleavage 3′ to the DSB sites releases SPO11 from DNA, leaving SPO11 covalently associated with an oligonucleotide. This chapter describes detection of the release product, SPO11-oligonucleotide complexes, from mouse testis lysates. The method for determining the size of SPO11-associated oligonucleotides is also provided.

Key words

SPO11 mouse testes terminal transferase DNA double-strand break 

References

  1. 1.
    Bellve, A.R. (1993) Purification, culture, and fractionation of spermatogenic cells. Methods Enzymol. 225, 84–113.PubMedCrossRefGoogle Scholar
  2. 2.
    Neale, M.J., Pan, J., Keeney, S. (2005) Endonucleolytic processing of covalent protein-linked DNA double-strand breaks. Nature 436, 1053–1057.PubMedCrossRefGoogle Scholar
  3. 3.
    Baudat, F., de Massy, B. (2005) Regulating double-stranded DNA break repair towards crossover or non-crossover during mammalian meiosis. Chromosome Res. 15, 565–77.CrossRefGoogle Scholar
  4. 4.
    Keeney, S., Baudat, F., Angeles, M., Zhou, Z.H., Copeland, N.G., Jenkins, N.A., Manova, K., Jasin, M. (1999) A mouse homolog of the Saccharomyces cerevisiae meiotic recombination DNA transesterase Spo11p. Genomics 61, 170–182.PubMedCrossRefGoogle Scholar
  5. 5.
    Romanienko, P.J., Camerini-Otero, R.D. (1999) Cloning, characterization, and localization of mouse and human SPO11. Genomics 61, 156–169.PubMedCrossRefGoogle Scholar
  6. 6.
    Sambrook, J., Russel, D.W. (2001) Molecular Cloning. Chapter 12. Protocols 8, 11, and 12. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York.Google Scholar

Copyright information

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

Authors and Affiliations

  • Jing Pan
    • 1
  • Scott Keeney
    • 2
  1. 1.Molecular Biology ProgramMemorial Sloan-Kettering Cancer CenterNew YorkUSA
  2. 2.Howard Hughes Medical Institute and Molecular Biology Program, Memorial Sloan-Kettering Cancer CenterNew YorkUSA

Personalised recommendations