page_title: Agarose gel electrophoresis
section_title: Electrophoresis
hierarchical_section_title: Agarose gel electrophoresis / General procedure / Electrophoresis
caption_attribution_description: Electrophoresis. Moving along gel. About halfway through.
context_page_description: Agarose gel electrophoresis is a method of gel electrophoresis used in biochemistry, molecular biology, genetics, and clinical chemistry to separate a mixed population of macromolecules such as DNA or proteins in a matrix of agarose, one of the two main components of agar. The proteins may be separated by charge and/or size, and the DNA and RNA fragments by length. Biomolecules are separated by applying an electric field to move the charged molecules through an agarose matrix, and the biomolecules are separated by size in the agarose gel matrix.
  Agarose gel is easy to cast, has relatively fewer charged groups, and is particularly suitable for separating DNA of size range most often encountered in laboratories, which accounts for the popularity of its use. The separated DNA may be viewed with stain, most commonly under UV light, and the DNA fragments can be extracted from the gel with relative ease. Most agarose gels used are between 0.7–2% dissolved in a suitable electrophoresis buffer.
context_section_description: Agarose gel electrophoresis is most commonly done horizontally in a submarine mode whereby the slab gel is completely submerged in buffer during electrophoresis. It is also possible, but less common, to perform the electrophoresis vertically, as well as horizontally with the gel raised on agarose legs using an appropriate apparatus.  The buffer used in the gel is the same as the running buffer in the electrophoresis tank, which is why electrophoresis in the submarine mode is possible with agarose gel.
  For optimal resolution of DNA greater than 2 kb in size in standard gel electrophoresis, 5 to 8 V/cm is recommended (the distance in cm refers to the distance between electrodes, therefore this recommended voltage would be 5 to 8 multiplied by the distance between the electrodes in cm).   Voltage may also be limited by the fact that it heats the gel and may cause the gel to melt if it is run at high voltage for a prolonged period, especially if the gel used is LMP agarose gel. Too high a voltage may also reduce resolution, as well as causing band streaking for large DNA molecules. Too low a voltage may lead to broadening of band for small DNA fragments due to dispersion and diffusion.
  Since DNA is not visible in natural light, the progress of the electrophoresis is monitored using colored dyes. Xylene cyanol (light blue color) comigrates large DNA fragments, while Bromophenol blue (dark blue) comigrates with the smaller fragments. Less commonly used dyes include Cresol Red and Orange G which migrate ahead of bromophenol blue. A DNA marker is also run together for the estimation of the molecular weight of the DNA fragments. Note however that the size of a circular DNA like plasmids cannot be accurately gauged using standard markers unless it has been linearized by restriction digest, alternatively a supercoiled DNA marker may be used.
caption: Agarose gel slab in electrophoresis tank with bands of dyes indicating progress of the electrophoresis. The DNA moves towards anode.
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