Concentrating DNA Samples
DNA samples are concentrated by precipitating them and then taking the sample up in a smaller volume of liquid. This is usually accomplished by using a combination of salt and alcohol. A common procedure is to use 0.3 M sodium acetate followed by ethanol. When precipitating small amounts of DNA, it’s important to carefully wash the sides of the tube to recover DNA that did not pellet to the bottom.
SUPPLIES NEEDED:
- 3 M Sodium acetate solution
- ethanol, absolute, ice cold
PROCEDURE:
1. Determine the volume of DNA that you have in your sample.
2. Calculate the amount of 3M sodium acetate that must be added to get a final concentration of 0.3 M.
A. This is done by using the formula X / (V + X) = C2 / C1
C1 = the starting molarity of the NaAc stock = 3M
C2 = the ending molarity of the NaAc stock = 0.3 M
X is the amount of NaAc to be added
V = the starting volume of DNA
For Example: You have 180 m L DNA (V)
The amount of NaAc to add (X) is:
X / (180 m l + X) = 0.3 / 3.0 = 1/10
10 X = 180 m L + X
9X = 180 m L
X = 20 m L of NaAc
B. Add this volume of 3 M NaAc to your DNA
C. Add 2 volumes of ice cold 100 % ETOH (freezer)
D. Precipitate for at least 30 mins in the freezer
E. Spin for 10 mins.
F. Pour off Ethanol.
G. Wash the sides of the tubes with 70% ETOH (freezer)
H. Spin again
I. Pour off ETOH
J. Dry under vacuum
K. Take up in TE