Agarose gel loading buffer

From OpenWetWare
Revision as of 04:21, 20 April 2007 by Jakob Suckale (talk | contribs) (Colour: table style)
Jump to: navigation, search

Loading dye is mixed with DNA samples for use in agarose gel electrophoresis. It generally contains a dye to assess how "fast" your gel is running and a reagent to render your samples denser than the running buffer (so that the samples sink in the well).



  • Ficoll
  • sucrose


dye 0.5-1.5% agarose 2.0-3.0% agarose* order # example
xylene cyanol 4000-5000 bp 750 bp Sigma X4126
bromophenol blue 400-500 bp 100 bp Sigma B8026
Orange G <100 bp  ?  ?

*sieving agarose


The precise amount of dye is not important. However, it is crucial that you prevent overlay of dye and expected DNA size. For example, if you are expecting a genotyping band of 200-400nt, you shouldn't use bromophenol blue since it will obscure your product. In this case, you should use a larger dye like xylene cyanol.

Ficoll & Orange G

  • Ficoll 400
  • Deionized water
  • Orange G dye

Dissolve 1.5 g of Ficoll in 10 mL of deionized water. Add very small amounts of Orange G dye such that the loading dye is dark orange. Store in small aliquots at 4°C (room temperature is okay too). To use, add and mix 1/5th volume of loading dye to DNA solutions prior to loading into the wells of gels.

Sucrose & xylene cyanol / bromophenol blue

  • 25mg bromophenol blue or xylene cyanol
  • 4g sucrose
  • H2O to 10mL

Store at 4°C to avoid mould growing in the sucrose. 10mL of loading buffer will last for years.

Specific recipes