PCR

• • allows scientists to rapidly amplify relatively small numbers of a piece of DNA millions of times - DNA is then suitable for gel electrophoresis and DNA profiling
  • used to diagnose disease, clone and sequence genes
• Requirements for PCR

  • template DNA
  • Taq polymerase
  • buffer solution to maintain pH
  • supply of 4 nucleotides (A, T, C, G)
  • 2 sets of primers
  • thermocycler
• Steps of PCR

  1. Denaturation (95°)
  2. Annealing (50-60°)
  3. Extension/Elongation (72°)
• Denaturation (95°)

  • double-stranded DNA melts open as weak hydrogen bonds are broken, creating 2 pieces of single-stranded DNA
  • all enzymatic reactions stop
• Annealing (50-60°)

  • primers bond to the beginning and end of the segment
  • primers anneal with the template strand according to the complementary base pair rule
  • attach to the 3' end
  • primers: short, single-stranded DNA molecules used to mark the 2 ends of a target sequence - tells polymerase enzymes where to start extending
• Extension/Elongation (72°)

  • Taq polymerase copies and extends DNA strand rapidly by joining complementary bases to the template strand - results in 2 copies of each original strand of the double-stranded DNA
  • DNA has doubles
  • Taq polymerase: enzyme that's able to resist denaturation at high temperatures - catalyses the formation of new DNA molecules from free nucleotides