Difficulties with DNA 1. 1.One cell normally provides too little material for study Gene cloning Polymerase Chain Reaction (PCR) 2. 2.There are often thousands of genes on a DNA molecule Electrophoresis

Restriction Enzymes Bacterial defense against viral DNABacterial defense against viral DNA Excise DNA at specific sequencesExcise DNA at specific sequences CCTTTG AATTCCCAGAATC GGAAACTTAA GGGTCTTAG AATTCGGCCATATACG GCCGGTATATGCTTAA Desired Gene Target Sites for EcoRI

Gene Cloning Recombinant plasmid Gene of interest Recombinant bacterium

Polymerase Chain Reaction In vitro amplification of a select length of DNAIn vitro amplification of a select length of DNA Denaturation Priming Elongation Desired Gene

Electrophoresis Separation of molecules based on sizeSeparation of molecules based on size Negatively charged DNA molecules are pulled through a gel by an electrical fieldNegatively charged DNA molecules are pulled through a gel by an electrical field Smaller molecules travel faster and fartherSmaller molecules travel faster and farther

Walter Jordan Poop

Restriction Fragment Length PolymorphismsAAGAATTCCCTGATCCATATATATATATCGGATCTAGAATTCGATTTCTTAAGGGACTAGGTATATATATATAGCCTAGATCTTAAGCTAAAGAATTCCCTGATCCATATATCGGATCTAGAATTCGATTGACTGTTCTTAAGGGACTAGGTATATAGCCTAGATCTTAAGCTAACTGACAAGAATTCCCTGATCCATATATCGGATCTAGAATTCGATTGACTGTTCTTAAGGGACTAGGTATATAGCCTAGATCTTAAGCTAACTGACAAGAATTCCCTGATCCATATATATCGGATCTAGAATTCGATTGACTTCTTAAGGGACTAGGTATATATAGCCTAGATCTTAAGCTAACTG Variations in DNA Variations in fragment sizes Variations in electrophoresis bands A B A B

Restriction Mapping (kilobases) Uncut plasmid Cut with EcoRI Cut with BamH3 Cut with Both DNA Marker