Agarose or gel electrophoresis is a molecular tool used in biology and biochemistry by research scientists to distinguish molecules, specifically nucleic acids like DNA or RNA, and proteins, of different sizes. It works on the principle that molecules of different sizes travel at different speeds through a gel, and therefore become separate from others. It is used as an analytical or diagnostic tool as a preliminary experiment for more complicated gene cloning or analysis projects.
Advantage Of Agarose
Agarose is relatively inexpensive compared to some other raw materials used in a research laboratory or in electrophoresis, such as polyacrylamide. It is also non-toxic before it is prepared and so easy to store on any laboratory bench or cabinet. During the gel-casting step of electrophoresis, it is easy to melt agarose in running buffers such as Tris-Borate-EDTA, and the suspension is easy to pour. In addition, it does not cause the sample to denature, so at the end of the electrophoresis procedure, precious or limited samples can still be recovered and used for other experimental procedures. If the gel is not required immediately, it can be placed in a plastic container and stored at 4 degrees C in a standard laboratory refrigerator for later use. Agarose gels have moderate to high resolution, depending on how they are prepared. The alternative to agarose is acrylamide, which must be cross-linked in order to form a solid gel, then becomes a neurotoxin.
Advantage of Agarose Electrophoresis
Since agarose melts easily in a standard microwave, it is easy to prepare or "pour" as a gel. Agarose electrophoresis is a visual method of confirming the presence of DNA, RNA, proteins, or a specific size(s) of these, unlike other detection methods, which give the precise quantities but not their size. It can give very good resolution and separation of large from small molecules as the agarose gel's pore size can be specified by the user, and only requires small amounts of sample. The process is fast (minimum of 30 minutes), and the apparatus is easy to set up and operate.
Disadvantages of Agarose Electrophoresis
Agarose gels may melt during the electrophoresis procedure because of high temperatures generated by the apparatus, which can also cause the buffer to evaporate and expose the gel. It is also sometimes hard to get different types of genetic molecules to run evenly, which may ruin the gel or make it difficult to recover the molecules for subsequent experiments without contamination by unwanted molecules. Also, if these molecules are very similar in size, then it may be difficult or impossible to resolve or separate them from each other. Quantification of the amount of a molecule by observing the size or intensity of fluorescence of a band may be difficult.
- "Gel Electrophoresis: Nucleic Acids: Essential Techniques;" P Jones, 1996
- Molecular Station: Agarose Gel Electrophoresis
- "Molecular Cloning: A Laboratory Manual;"Sambrook J, Russel DW; 2001
- University of Illinois: Gel electrophoresis of DNA
- Queen's University: Agarose Gel Electrophoresis