Agarose gel electrophoresis

Contributed by Ivan Delgado <>

The preparation, and running, of an agarose gel for nucleic acid separation.


10X DNA loading buffer (25% Ficoll (Type-400) in dH2O (note: 25% Ficoll is dense and will not go into solution readily, just stir for about 1-2 hours). The buffer is ready for use. But it is a very good idea to add a dye to not only be able to see your sample as you load it on a gel, but to track what size fragments during the run. Any combination of these dyes can be used: 0.25% Bromo-Phenol-blue (300 bp); 0.25% Xylene Cyanol (1-2 kb); 0.25% Orage-G (50 bp). Also, EtBr can be added to the buffer or the gel at a concentration of ~1 mL/10 mL of EtBr. I find it to be better to add it directly to the gel to prevent degradation during storage of the buffer) 5X TBE (Mix 54 g of Tris base, 27.5 g of boric acid, 20 mL of 0.5M EDTA, pH 8.0, and dH2O to 1 L) 1X TBE (Mix 100 mL of 5X TBE with 900 mL of dH2O)


  • The concentration of agarose needed to resolve the following fragment sizes:

1.2% = 100 bp - 5 kb 2% = 100 bp - 2 kb 4% = 20 bp - 500 bp

  • Mix the desired amoune of agarose with 1X TBE in a flask. For a 1% gel, add 1 g of agarose to 100 mL of 1X TBE
  • Microwave into solution (while microwaving, take flask out of microwave swirl a few times). It is important the the agarose is completely into solution
  • Add EtBr if desired

RECOMMENDED: add EtBr to the gel, this way it is quicker to visualize the DNA fragments right after the gel run

  • Pour gel into agarose gel set up
  • Wait until the gel solidified (~1 hr)
  • Mix DNA samples with loading buffer (1 mL of 10X loading buffer for each 9 mL of DNA sample)
  • Carefully load DNA samples into the wells of the agarose gel
  • Using 1X TBE as running buffer, run the agarose gel (100 V is typically more than enough)
  • Visualize the DNA bands on a UV box or Immaging system

This method is based, with permission, on an original protocol available here.