UNIT 2.9A Southern Blotting

Southern blotting is the transfer of DNA fragments from an electrophoresis gel to a membrane support (the properties and advantages of the different types of membrane, transfer buffer, and transfer method are discussed in detail), resulting in immobilization of the DNA fragments, so the membrane carries a semipermanent reproduction of the banding pattern of the gel. After immobilization, the DNA can be subjected to hybridization analysis, enabling bands with sequence similarity to a labeled probe to be identified. This unit presents several protocols for DNA transfer to nylon and nitrocellulose membranes, and for calibrating a UV transilluminator to cross-link the transferred DNA to the membrane.