AGAROSE GEL ELECTROPHORESIS (AGE) Theory before performing electrophoresis

AGAROSE GEL ELECTROPHORESIS (AGE)

Theory before performing electrophoresis
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       AGE is the most convenient and easiest way to separate DNA molecules. The DNA molecules are separated on the basis of their size under the influence of an electric charge applied to the electrophoretic apparatus. The smaller molecules will travel faster and farther as compared to the bigger molecules which travel slower.

Agarose gel is obtained from Red Algae (Porphyra umbilicalis). The IUPAC name of agarose gel is

(1 4)-3,6-anhydro – α – L – galactosepyranosyl – (1 3) – β – D – galactopyranan

In general agarose gel is prepared between 0.2 – 0.7%. The 0.7% gel is ideal for the separation of larger molecules. And the 0.2% gel is ideal for separation of smaller molecules.

EAUIPMENTS AND MATERIAL NECESSARY FOR AGE:

1.       Electrophoretic apparatus:

2.       Gel casting trays:

3.       Sample combs: to form wells for samples to be loaded in the gel.

4.       Loading buffer: it allows the sample to be filled in the wells.

5.       Tracking dyes: it migrates in the gel with the DNA fragments and provides visualization for the proceeding status of the electrophoresis.

6.       Ethidium bromide: it is a fluorescent dye which can be seen under ultraviolet rays. It incorporates with the DNA fragments and when the gel is visualized under the UV rays. The ethidium bromide fluoresces to mark the location of the travelled DNA fragments in the gel.

7.       Transilluminator: it is an ultraviolet box. Which is used to visualized ethidium bromide stained separated DNA fragments after electrophoresis..