Biology Protocols

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Agarose gel electrophoresis

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Agarose gel electrophoresis is the easiest and most common way of separating and analyzing DNA. An example apparatus is shown in the figure below.

 

 

The purpose of the gel might be to look at the DNA size, to quantify it or to isolate a particular band. The DNA is visualised in the gel by addition of ethidium bromide. This binds strongly to DNA by intercalating between the bases and is fluorescent meaning that it absorbs invisible UV light and transmits the energy as visible orange light.

The principal of the technique is that DNA, a negatively charged molecule, will migrate through a predetermined agarose matrix with an electric field towards the anode (positive electrode). Migration though the agarose matrix is easier for smaller DNA molecules and more difficult for larger DNA molecules meaning that different sized molecules can be separated.

DNA samples are run against a DNA ladder comprising DNA fragments of known sizes, the most common of which is the 1kb marker (see below).

 


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