Electroelution of DNA from Agarose

Electroelution of agarose fragments

Electroelution buffer

1 M Tris, pH 7.5 12.0 mls

0.5 M EDTA 0.24 mls

1 M NaCl 3.0 mls

qs to 600 mls dH2O

Acetate cushion

3 M NaAcetate pH 4.8 480 μl

0.1 % Bromphenol Blue 40 μl

1. Place gel slices in trough

2. Remove all air bubbles, then layer 80 μl of acetate cushion

3. Electroelute at: 120V for ~1Kb to 140V for >2.5Kb

for 40 min for ~1Kb to 60 min for >2.5Kb

4. Collect ~300 μl of salt cushion, add 3X volumes of 95% ethanol to precipitate

5. Remove gel slices

Clean wells

Run for 10 min longer

Clean wells again

Rinse thoroughtly to remove any extraneous DNA