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Gel electrophoresis is an electrophoresis method for separation and analysis of biomacromolecules (DNA, RNA, proteins, etc.) and their fragments, based on their size and charge through a supportive medium.
Electrophoresis techniques used in the assessment of DNA damage include alkaline gel electrophoresis and pulsed field gel electrophoresis. For short DNA segments such as 20 to 60 bp double stranded DNA, running them in polyacrylamide gel (PAGE) will give better resolution (native condition). [ 1 ]
Two-dimensional gel electrophoresis, abbreviated as 2-DE or 2-D electrophoresis, is a form of gel electrophoresis commonly used to analyze proteins. Mixtures of proteins are separated by two properties in two dimensions on 2D gels. 2-DE was first independently introduced by O'Farrell [ 1 ] and Klose [ 2 ] in 1975.
The purpose of gel electrophoresis is to separate proteins by physical or chemical properties, which include charge, molecular size, and pH.< When separating based on size, the ideal method is SDS-PAGE or polyacrylamide gel electrophoresis and molecular-weight size markers are the appropriate standards to use.
The limit of resolution for standard agarose gel electrophoresis is around 750 kb, but resolution of over 6 Mb is possible with pulsed field gel electrophoresis (PFGE). [7] It can also be used to separate large proteins, and it is the preferred matrix for the gel electrophoresis of particles with effective radii larger than 5–10 nm.
During electrophoresis in a discontinuous gel system, an ion gradient is formed in the early stage of electrophoresis that causes all of the proteins to focus into a single sharp band. The formation of the ion gradient is achieved by choosing a pH value at which the ions of the buffer are only moderately charged compared to the SDS-coated proteins.
Difference gel electrophoresis (DIGE) is a form of gel electrophoresis where up to three different protein samples can be labeled with size-matched, charge-matched spectrally resolvable fluorescent dyes (for example Cy3, Cy5, Cy2) prior to two dimensional gel electrophoresis.
Pulsed-field gel electrophoresis (PFGE) is a technique used for the separation of large DNA molecules by applying an electric field that periodically changes direction to a gel matrix. [ 1 ] [ 2 ] Unlike standard agarose gel electrophoresis , which can separate DNA fragments of up to 50 kb, PFGE resolves fragments up to 10 Mb. [ 1 ]
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