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Protocol for the Taq Control PCR Reaction

Introduction

Amplification of a 500 bp Lambda Fragment from the control mix. All kit components should be mixed and spun down prior to pipetting.

Protocol

  1. Prepare the following 50 µl reaction in a 0.5 ml PCR tube on ice:
    38.8 µl nuclease-free water
    5 µl 10X Standard Taq Reaction Buffer
    1 µl 10 mM dNTP Solution Mix
    5 µl Control Template/Primer Mix
    0.2 µl Taq DNA Polymerase*
    * Due to the difficulties in pipetting small volumes of enzyme, Taq DNA Polymerase can be diluted in Diluent F (NEB #B8006S) or 1X reaction buffer. For example, 1 µl of Taq DNA Polymerase is mixed with 4 µl of diluent and 1 µl of that mixture is added to the reaction. Enzyme diluted in Diluent F can be stored at -20°C for future use.
  2. Gently mix the reaction and spin down in microcentrifuge.
    If the thermocycler does not have a heated cover, add one drop of mineral oil to the reaction tube to prevent evaporation.
  3. Cycling conditions for Control PCR: