Southern, Western and Northern Blotting

Abstract

Southern blotting, discovered in 1975 by E.M. Southern, represents a technique to detect a gene of interest in the DNA sample. The steps involved are isolation of DNA, its separation by electrophoresis, transfer to a suitable medium, hybridization to probes and visualization of the gene if it is present. Western blotting is the counterpart which is used to detect proteins. The difference lies in the visualization process. In Western blotting, this is made possible by primary and secondary antibodies, whereas in Southern blotting, a radiolabeled (fluorescent) probe or dye that binds to the DNA is used. Application of Western blotting includes identifying HIV antigens or Hepatitis B surface antigen in blood. Northern blotting is used to detect mRNA of interest, where after separation by electrophoresis, cDNA is used as a probe that binds to the RNA strand; the application includes finding alternate transcript size. The term ‘blotting’ in all the three techniques represents the transfer of material after separation to nitrocellulose paper by means of diffusion.