Ethanol precipitation: Difference between revisions
Plantsurfer (talk | contribs) ethanol precipitation is also used to precipitate polysaccharides from aqueous solutions |
Enemyunknown (talk | contribs) Added 2 links to good publications on ethanol precipitation |
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==References== |
==References== |
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{{Unreferenced|date=December 2007}}<references/> |
{{Unreferenced|date=December 2007}}<references/> |
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==External links== |
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* {{cite journal |author=Zeugin JA, Hartley JL |title=Ethanol Precipitation of DNA |journal=Focus |volume=7 |issue=4 |pages=1–2 |year=1985 |url=http://www.invitrogen.com/etc/medialib/en/filelibrary/pdf/focus.Par.56415.File.dat/Focus%20Volume%207%20Issue%204.PDF |accessdate=2008-09-10}} |
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* {{cite journal |author=Crouse J, Amorese D |title=Ethanol Precipitation: Ammonium Acetate as an Alternative to Sodium Acetate|journal=Focus |volume=9 |issue=2 |pages=3–5 |year=1987|url=http://www.invitrogen.com/etc/medialib/en/filelibrary/pdf/focus.Par.34900.File.dat/Focus%20Voume%209%20Issue%202.pdf |accessdate=2008-09-10 }} |
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[[Category:Laboratory techniques]] |
[[Category:Laboratory techniques]] |
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Revision as of 18:21, 10 September 2008
Ethanol precipitation is a method used to concentrate DNA and polysaccharides such as pectin and xyloglucan from solution in water. DNA is polar and soluble in water because water is polar. Based on the principle of "like dissolves like", it is insoluble in the relatively nonpolar ethanol.
DNA is initially precipitated by adding a large volume of 100% ethanol, and cooling on ice or dry ice. This will precipitate DNA, as well as the salts that form ionic bonds with it. The suspension is centrifuged in a microcentrifuge tube at high speeds (~12,000g), and the supernatant solution removed, leaving a pellet of the crude DNA. The DNA precipitation in this step is due to the ethanol interacting with the water such that fewer water molecules are available to dissolve the DNA molecules.
In the next step, 70%-80% ethanol is added to the precipitated pellet, and is gently mixed to resuspend the DNA. This allows the 20-30% water to access the salts present in the pellet. This suspension is centrifuged again, and the supernatant solution is removed. This step is repeated once.
Finally, the pellet is air-dried giving the purified DNA.
Isopropanol can be used instead of ethanol; the precipitation efficiency of the isopropanol is higher. However, isopropanol is less volatile than ethanol and needs more time to air-dry in the final step.
Method
- Add 1/10 volume of Sodium Acetate (3M, pH 5.2).
- Add 2.5-3.0X volume of 100% ethanol.
- Incubate at -80oC for 15 minutes.
- Centrifuge at > 14,000 x g for 30 minutes at 4 degrees Celsius.
- Rinse with 70% Ethanol
See also
References
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External links
- Zeugin JA, Hartley JL (1985). "Ethanol Precipitation of DNA" (PDF). Focus. 7 (4): 1–2. Retrieved 2008-09-10.
- Crouse J, Amorese D (1987). "Ethanol Precipitation: Ammonium Acetate as an Alternative to Sodium Acetate" (PDF). Focus. 9 (2): 3–5. Retrieved 2008-09-10.