Electrophoresis separates charged particles, commonly used in biology to separate DNA or protein molecules. Various procedures are used based on the type and size of molecules. Gels are commonly used as a support medium, with agarose for DNA and acrylamide for proteins. Capillary and 2-D electrophoresis are advanced techniques.
The electrophoresis process uses an electric charge field to separate charged particles. It is commonly used in biology to separate DNA or protein molecules. Various procedures can be used, depending on the type and size of the molecules to be separated, but all procedures require a charge source, support medium, and buffer solution.
All electrophoresis procedures separate molecules based on their charge and size. An electric field is applied to the molecules, and since the molecules are electrically charged, a force acts on the molecules as a result. The higher the charge of the molecule, the greater the force applied by the electric field and the further the molecule will move through the support medium. Size also affects how far a molecule travels: a large molecule won’t move as far as a small molecule with the same charge. The ratio of charge to mass for a molecule determines how far it moves through the medium of support.
Various types of gels are the most commonly used support medium for electrophoresis. The gels can be in the form of plates or tubes. Gel plates allow you to run many samples at once, so they are the most commonly used method in laboratories. Tube gels, however, allow for better resolution of sample results, so they are sometimes a better choice for protein electrophoresis.
Agarose gel is a common substance used for electrophoresis of DNA and other nucleic acids. Agarose creates a large pore structure, so large molecules that often have to be passed through the gel to analyze DNA can move around more easily. However, a different type of gel is usually used if the goal is to sequencing smaller DNA molecules. This gel, called denaturing polyacrylamide gel or simply sequencing gel, provides results with much higher resolution. The results allow a scientist to distinguish between two segments of DNA that differ in only one base pair.
Acrylamide gels are generally used for protein separation. The most common process is called sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). SDS is a detergent that denatures proteins. The gel is run with a buffer that contains a low-mobility ion layer and a high-mobility ion layer. These layers help separate proteins based on their size. Sometimes the proteins are also run in native gels, which do not denature the proteins.
Two more advanced techniques are capillary and 2-D electrophoresis. The capillary procedure forces molecules through a capillary tube with electrical charge and provides highly accurate results. 2-D electrophoresis separates molecules along an x-axis and a y-axis. Molecules are separated by size along one axis and by charge along the other axis.
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