Southern Blotting

Southern blotting is the process of transferring and immobilizing DNA onto a membrane after restriction enzyme digestion and electrophoresis. The purpose is to subsequently probe the membrane with DNA to find complementary sequences. A complementary fragment of DNA is used to detect specific DNA sequences immobilized onto a membrane from a Southern blot. Two probes, DYZ1 and D2S44, are commonly used for identification of individuals. DYZ1 is a Y chromosome-specific probe, and D2S44 detects a restriction fragment length polymorphism (RFLP) on chromosome 2. The probe serves as a label or tag to identify specific regions of DNA out of the total 3 billion base pairs of DNA found in the human genome. Long, relatively intact pieces of DNA are required for restriction enzyme digestion for Southern blot analysis of human genomic DNA. Approximately 10- to 20-pg quantities are required to identify a single-copy region from the human genome. This section also presents an exercise on restriction fragment length polymorphism analysis of human DNA.