Preparation of Genomic DNA from Mammalian Tissue

William M. Strauss1

1 Harvard Medical School and Beth Israel Deaconess Medical Center, Boston, Massachusetts
Publication Name:  Current Protocols in Neuroscience
Unit Number:  Appendix 1H
DOI:  10.1002/0471142301.nsa01hs06
Online Posting Date:  May, 2001
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Abstract

There are a number of different procedures for the preparation of genomic DNA. They all start with some form of cell lysis, followed by deproteination and recovery of DNA. The main differences between various approaches lie in the extent of deproteination and in molecular weight of the DNA produced. The isolation procedure described here is relatively brief and relies on the powerful proteolytic activity of proteinase K combined with the denaturing ability of the ionic detergent SDS.

     
 
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Table of Contents

  • Preparation of Genomic DNA from Mammalian Tissue
  • Reagents and Solutions
  • Commentary
     
 
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Materials

Basic Protocol 1:

  Materials
  • Tissues, whole or cultured cells
  • Liquid nitrogen
  • Digestion buffer (see recipe)
  • PBS ( appendix 2A), ice cold
  • 25:24:1 (v/v/v) phenol/chloroform/isoamyl alcohol ( appendix 2A)
  • 7.5 M ammonium acetate
  • 70% and 100% ethanol
  • TE buffer, pH 8 ( appendix 2A)
  • Incubator or water bath at 50°C, with shaker
  • Additional reagents and equipment for trypsinizing adherent cells ( appendix 3B) and phenol/chloroform/isoamyl alcohol extraction ( appendix 1G)
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Figures

Videos

Literature Cited

Literature Cited
   Enrietto, P.J., Payne, L.N., and Hayman, M.J. 1983. A recovered avian myelocytomatosis virus that induces lymphomas in chickens. Pathogenic properties and their molecular basis. Cell 35:369‐379.
   Gross‐Bellard, M., Oudet, P., and Chambon, P. 1973. Isolation of high‐molecular‐weight DNA from mammalian cells. Eur. J. Biochem. 36:32‐38.
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