Tip: The depurination step should not last too long, since very short fragments attach less firmly to the membrane. Rinse the gel briefly in distilled water. During this period the color of the bromophenol blue in the samples will change from blue to yellow, indicating that the gel has been completely saturated with the acid.
In order to facilitate their transfer, these fragments are reduced in size, either by acid depurination or by UV irradiation.Īcid depurination - immediately after gel electrophoresis, place the gel in a solution of 0.2 M HCl, so that it is completely covered.
A standard protocol is described here together with recipes for buffers and solutions. Many variations on the Southern blotting procedure exist. The DNA of interest can be identified by hybridization to radioactive or chemiluminescent probes and visualized by autoradiography or staining. Southern) refers to the transfer of the DNA to a nylon or nitrocellulose membrane by capillary transfer. Southern blotting (named after its inventor, E.M. The DNA is next run through an agarose gel (6). DNA is usually first converted into conveniently sized fragments by restriction digestion. Southern blotting is a widely used technique that allows analysis of specific DNA sequences.
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