Precipitation DNA

Step ;
1. Adjust the concentration of monovalent cations in the sample by adding
1/10th volume of 3 M sodium acetate to the sample
2. Add 2 volumes of ethanol and mix gently. If there is sufficient DNA in the
sample, you will see a white precipitate form very rapidly
3. Recover the DNA by centrifugation, and rinse the resulting pellet with 70%
ethanol to remove residual salt
4. Evaporate off residual ethanol and resuspend the DNA in the desired buffer to
the desired concentration.
The image to the right shows samples of genomic DNA before and after addition
of sodium acetate and ethanol. The precipitate became visible within a few
seconds of adding ethanol. When the DNA in contained in small volumes, the
procedures is usually carried out in 1.5 ml microcentrifuge tubes.
Like many procedures in molecular biology, there are several effective
variations-on-a-theme for precipitating DNA from solution, and with a few
exceptions, the choice of which to use is largely a matter of personal preference.
The significant variables for nucleic acid precipitation are :
-

Type and concentration monovalent cation : the frequently-used sources of

cations are :
o Sodium acetate at a final concentration of 0.3 M (3M stock solution)
o Sodium chloride at a final concentration of 0.2 M (5M stock solution)
o Ammonium acetate at a final concentration of 2.5 M (7.5 M stock
solution)
o Lithium chloride at a final concentration of 0.8 M (8 M stock solution).
In some situations, one salt is preferred over another. For example,
ammonium acetate should not be used if the DNA is going to be
phosphorylated with polynucleotide kinase. If the DNA contains the
detergent SDS, sodium Chloride is the choice because it allows SDS to
remain soluble in 70% ethanol.

Ethanol versus isopropanol : isopropanol is an effective alternative to ethanol

and has the advantage of precipitating DNA at lower concentrations. Instead
of mixing 2 volumes of ethanol with the DNA-salt-solution, addition of one
volume of isopropanol will suffice.
Time and temperature allowed for precipitation : precipitation occurs very
rapidly except when DNA content is very low (i.e. <100 ng ). In most case,
you can start centrifugation immediately after adding ethanol or isopropanol.
Many older manuals indicate that precipitation should be allowed to occur in
the cold (e.g. -20C). this has been shown to be unnecessary. Typically,
centrifugations is at about 12,000 x g for 10-15 minutes.
Drying DNA before resuspension : the purpose here is to allow evaporation of
ethanol or isopropanol. It is best not to overdry the DNA, which makes

resolution more difficult and can cause denaturation of DNA. Drying is best
achieved by leaving the tube on the lab bench for a few minutes.