One of the fundamental methods in molecular biology is southern blotting. In Southern Blotting, DNA molecules, typically restriction fragments, are transferred from an electrophoresis gel to a nitrocellulose or nylon sheet (referred to as a "membrane") in order to replicate the DNA banding pattern that exists in the gel.
The analysis of DNA fragments within a specific size range and identification of those fragments based on their molecular weight is done using semi-automated Southern Blotting. DNA fragments are separated and identified using a combination of human and automated methods. The detection and analysis of genetic alterations, the presence of foreign DNA, and the size of DNA fragments can all be done using semi-automated Southern blotting.
The next step in Southern Blotting is the hybridization of the transferred DNA fragments with a labeled DNA probe. The probe is a single-stranded DNA molecule that is complementary to the target DNA sequence of interest. The probe is labeled with a detectable molecule, such as a radioactive isotope or a fluorescent dye, which allows the detection of the hybridized DNA fragment.
The final step in Southern Blotting is the detection of the specific DNA fragment of interest. The labeled probe hybridizes to the complementary DNA fragment on the membrane, forming a stable double-stranded molecule. The membrane is then washed to remove any unbound probe, and the labeled hybridized DNA fragment is detected using autoradiography, chemiluminescence, or fluorescence imaging
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