We will use agarose gel electrophoresis to determine DNA fragment sizes, and to quantify DNA.
This technique separates DNA molecules based on size. DNA has a net negative charge that is proportional to its size. When placed in a matrix of agarose and exposed to an electric field, DNA molecules will move towards the positive electrode. The agarose gel is made up of tiny holes. Smaller molecules move faster than lager molecules because they produce less friction in the agarose matrix.
Following electrophoresis the DNA is visualized and the gel is photographed. The distance each DNA fragment moves from the gel origin is directly proportional to the size of the DNA fragment. If a DNA ladder (a standard made up of several DNA molecules of known length) was run on the gel, we can estimate the sizes of the unknown DNA molecules. We can also use the DNA ladder to estimate the concentration of DNA molecules in a given solution.
Agarose gel electrophoresis has many variable that are application dependant. For example, you must decide
- what buffer to use
- the concentration of agarose to use
- how to prepare the DNA sample for loading
- the voltage and run time to use
- what DNA ladder to use
See the protocol “Agarose gel electrophoresis of DNA” to determine the correct parameters for your application.
We commonly refer to agarose gel electrophoresis as 'running a gel'.
Run a virtual gel