Introduction:

Routine use agarose is ideal for everyday analysis of nucleic acids by gel electrophoresis or blotting (Northern or Southern) and is also suitable for protein applications such as Ouchterlony and radial immunodiffusion (RID).

Analysis Note :

Sulfate content - used as an indicator of purity, since sulfate is the major ionic group present.
Gel strength - the force that must be applied to a gel to cause it to fracture.
Gel point - the temperature at which an aqueous agarose solution forms a gel as it cools. Agarose solutions exhibit hysteresis in the liquid-to-gel transition - that is, their gel point is not the same as their melting temperature.
Electroendosmosis (EEO) - a movement of liquid through the gel. Anionic groups in an agarose gel are affixed to the matrix and cannot move, but dissociable counter cations can migrate toward the cathode in the matrix, giving rise to EEO. Since electrophoretic movement of biopolymers is usually toward the anode, EEO can disrupt separations because of internal convection.

Preparation of Agarose Gels for DNA separations:

Weigh out the desired amount of agarose and place in an Erlenmeyer flask with a measured amount of electrophoresis buffer, e.g. for an 0.8% gel, add 0.8 gm of agarose and 100ml of TBE Buffer (1X), to a 200 ml flask. The larger flask insures against the agarose boiling over. Dissolve the agarose in a boiling water bath or in a revolving-plate microwave oven. All the grains of agarose should be dissolved and the solution clear. Cool the solution to 60°C ( 70°C for concentrations 2% or above) and pour immediately. Allow the gel to set for one-half hour before using. Make sure to use the same electrophoresis buffer in the gel as for the running buffer.