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PCR Master Mix Calculator

Calculate Q5 Master Mix Volumes with 10% Overage

Input Parameters

Include your samples + controls
Default: 25 μL per reaction

Q5 Components

Standard Q5 PCR reaction:

  • Q5 Buffer (5X): 20% of reaction
  • dNTPs (10 mM): 0.2 mM final
  • Forward Primer: 0.5 μM final
  • Reverse Primer: 0.5 μM final
  • Q5 Polymerase: 0.02 U/μL
  • DNA Template: 10 ng per reaction
  • Water: To final volume

Note: DNA template is added individually to each tube, NOT to master mix!

Master Mix Volumes (with 10% overage)

Component Volume per Reaction Total Volume

10% overage added to account for pipetting error and dead volume

Mix components in order listed, vortex gently, and aliquot into PCR tubes

Per-Reaction Breakdown

Component Volume per Tube (μL)
Master Mix -
DNA Template 10 μL
Total Reaction Volume -

Add - μL of master mix to each tube first

Then add 10 μL of DNA template (calculated from DNA Input Calculator)