PCR and Electrophoresis

Techniques of Recombinant DNA Technology Multiplying DNA in vitro: The Polymerase Chain Reaction (PCR) VIDEO CLIP ◦Large number of identical molecules of DNA produced in vitro ◦Critical to amplify DNA in variety of situations  Epidemiologists use to amplify genome of unknown pathogen  Amplified DNA from Bacillus anthracis spores in 2001 to identify source of spores 2

Techniques of Recombinant DNA Technology Multiplying DNA in vitro: The Polymerase Chain Reaction (PCR ) ◦Repetitive process consisting of three steps  Denaturation  Priming  Extension ◦Can be automated using a thermocycler 3

Polymerase chain reaction (PCR) 4 Figure 8.5a

Polymerase chain reaction (PCR) 5 Figure 8.5b

Techniques of Recombinant DNA Technology 6

Separating DNA Molecules: Gel Electrophoresis and the Southern Blot ◦Gel electrophoresis  Separates molecules based on electrical charge, size, and shape  Allows scientists to isolate DNA of interest  Negatively charged DNA drawn toward positive electrode  Agarose makes up gel; acts as molecular sieve  Smaller fragments migrate faster and farther than larger ones  Determine size by comparing distance migrated to standards 7

Gel electrophoresis 8 Figure 8.6

GEL ELECTROPHORESIS VIDEO Video of our lab’s gel electrophoresis Electrophoresis Results video 9