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Methods:DNase I nicking

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This method is used to release the constraints of supercoiling while keep the other topologies of substrate DNA (i.e. catenanes, knots, etc.)

Materials Needed

According to Colloms et al [1], the reaction mix is:

  • 50mM Tris-HCl (pH 8.0)
  • 10 mM MgCl2
  • 50 mM NaCl
  • 0.3 mg/ml EtBr
  • 1 µg/ml DNase I

According to Peng et al[2], the reaction mix is:

  • 40mM Tris-HCl (pH 7.6 at 30C)
  • 6 mM MgCl2
  • 20 mM KCl
  • 2 mM DTT
  • 1-8ng DNase I (various amounts were used in paper)


According to Peng et al [2]:

Total reaction volume: 20µl
Incubation Time

  • 30C for 30 seconds

Termination of the reaction:

  • 20µl of stop solution

Product Recovery

  • Phenol extraction/ EtOH precipitation


User notes


See Help:References for how to manage references in EcoliWiki.

  1. Colloms, SD et al. (1997) Topological selectivity in Xer site-specific recombination. Cell 88 855-64 PubMed EcoliWiki page
  2. 2.0 2.1 Peng, H & Marians, KJ (1995) The interaction of Escherichia coli topoisomerase IV with DNA. J. Biol. Chem. 270 25286-90 PubMed EcoliWiki page