IGEM:Harvard/2006/TOPO Cloning

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  • After purifying the PCR product, add Taq polymerase buffer and dATP to their working concentrations, and 0.5 unit of Taq polymerase. Incubate the reaction for 10-15 minutes at 72°C.
  • To prepare the cloning reaction combine the following:
    • 0.5 to 4 uL PCR product
    • 1 uL salt solution
    • dilute to a total of 5 uL with water
    • 1 uL TOPO vector
  • Mix gently and incubate at room temperature for 5 minutes.
  • Put the reaction on ice and proceed to the standard TOP10 transformation protocol.