Abstract
Extraction procedures for plant DNA in general must accomplish the following.
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(1)
The cell walls must be broken (or digested away) in order to release the cellular constituents. This is usually done by grinding the tissue in dry ice or liquid nitrogen with a mortar and pestel or a food grinder.
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(2)
The cell membranes must be disrupted, so that the DNA is released into the extraction buffer. This is accomplished by using a detergent, usually SDS (sodium dodecyl sulfate) or CTAB (cetyltrimethylammonium bromide).
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(3)
The DNA must be protected from the endogenous nucleases. The detergents are used for this purpose, as is EDTA (ethylenediaminetetraacetic acid). It is a chelating agent that binds magnesium ions, generally considered a necessary cofactor for most nucleases (but see note f, below). In addition, the buffer/tissue mixture is emulsified with either chloroform or phenol to denature and separate the proteins from the DNA.
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(4)
Shearing of the DNA should be minimized. DNA in solution can be broken by exposure to turbulence (e.g., being quickly drawn through a small orifice). Typically, DNA 50–100 kb in length can be obtained without great care being taken.
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(5)
The time between thawing of the frozen, pulverized tissue and its exposure to the extraction buffer should be minimized to avoid nucleolytic degradation of the DNA.
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© 1989 Kluwer Academic Publishers, Dordrecht
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Rogers, S.O., Bendich, A.J. (1989). Extraction of DNA from plant tissues. In: Gelvin, S.B., Schilperoort, R.A., Verma, D.P.S. (eds) Plant Molecular Biology Manual. Springer, Dordrecht. https://doi.org/10.1007/978-94-009-0951-9_6
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DOI: https://doi.org/10.1007/978-94-009-0951-9_6
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