How to Make an Agarose Gel

Choose the percentage

The percentage sets the size of DNA the gel separates best. Lower percentages leave larger pores for large fragments; higher percentages resolve small ones.

• 0.7%: about 0.8 to 12 kb
• 1%: about 0.5 to 10 kb, the usual choice for PCR products and plasmids
• 1.5%: about 0.2 to 3 kb
• 2%: about 50 bp to 2 kb

Work out the amounts

A gel percentage is grams of agarose per 100 mL of buffer: grams = % × volume (mL) ÷ 100. A 50 mL, 1% gel needs 0.5 g. Make the buffer at 1× from a stock: 1 mL of 50× TAE per 50 mL, or 5 mL of 10× TBE. Use the same buffer in the tank.

Pour the gel

1. Weigh the agarose into a flask and add the 1× buffer.
2. Heat in a microwave in short bursts, swirling between them, until the solution is completely clear with no floating grains. Watch it closely: it can boil over suddenly.
3. Let it cool to about 50 to 60 °C, cool enough to hold the flask.
4. Add DNA stain if you stain in the gel, swirl, and pour into the tray with the comb in place.
5. Leave it to set for 20 to 30 minutes, then remove the comb carefully.

Run it

Place the gel in the tank with the wells at the negative (black) electrode, since DNA runs toward the positive (red) one, and cover it with buffer. Load the samples mixed with loading dye alongside a DNA ladder, and run at about 5 to 8 volts per cm between the electrodes until the dye front is two-thirds of the way down.

Calculators

The Agarose Gel Calculator gives the agarose, buffer, water and stain for any percentage and volume. To load a set amount of DNA, measure it first with the DNA Concentration Calculator.