Team:Calgary/Notebook/Protocols/KapaPCR

From 2013.igem.org

Kapa PCR (for DNA constructions)

Mix (per reaction)

  • 5.0µL 5x Kapa Hifi Buffer
  • 0.75µL 10mM dNTPs
  • 0.75µL Primer Forward
  • 0.75µL Primer Reverse
  • 0.5µL Kapa Hifi DNA polymerase
  • 10 to 50ng of DNA
  • ddH2O (deionized water) up to 25µL

Protocol

  1. Add the mix to each tube
  2. Set the following program in the thermocycler:
    1. Stage 1 (1 cycle): 95°C for 2 to 5 min
    2. Stage 2 (30 cycles): 98°C for 20 sec, 55°C for 15 sec, 72°C for 1min/kb
    3. Stage 3 (1 cycle): 72°C for 1 to 5 min
    4. Hold at 4°C
  3. Run the products in a agarose gel