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Master Mix Calculator

Scale a reaction recipe into one master mix: per-reaction and batch volumes for every component, extra for pipetting loss, and the water that makes up the volume.

Formula

Vrxn,i=Vrxn×Cfinal,iCstock,iV_{\text{rxn},i} = V_{\text{rxn}} \times \dfrac{C_{\text{final},i}}{C_{\text{stock},i}}
Vwater=Vrxn−Vsample−∑VotherV_{\text{water}} = V_{\text{rxn}} - V_{\text{sample}} - \sum V_{\text{other}}
Vmix,i=Vrxn,i×Nrxn×(1+extra100)V_{\text{mix},i} = V_{\text{rxn},i} \times N_{\text{rxn}} \times \left(1 + \dfrac{\text{extra}}{100}\right)
VrxnV_{\text{rxn}}
total volume in each tube, including the sample
VsampleV_{\text{sample}}
the volume added to each tube separately, outside the master mix
extra\text{extra}
overage that covers liquid lost in tips and on tube walls

How it works

A master mix combines everything the reactions share, so each tube receives one accurate transfer instead of several small ones. Every reaction then has the same composition, and the differences between tubes come from the samples alone.

Components with a stock and a final concentration are scaled by C₁V₁ = C₂V₂ against the reaction volume. Components dosed by volume, such as an enzyme added at 1 µL per reaction, are entered directly. Water takes up whatever volume is left once the sample is accounted for.

Worked example

Ten 20 µL reactions with 10% extra: 5× buffer, 10 mM dNTPs to 0.5 mM, 10 µM primer to 0.5 µM, 1 µL of enzyme and 2 µL of sample each.

  1. Buffer: 20 µL × 1 / 5 = 4 µL. dNTPs: 20 µL × 0.5 / 10 = 1 µL. Primer: 20 µL × 0.5 / 10 = 1 µL.
  2. Water = 20 − 2 − 4 − 1 − 1 − 1 = 11 µL.
  3. Scale by 10 reactions × 1.1 = 11.

Mix 121 µL of water, 44 µL of buffer and 11 µL each of dNTPs, primer and enzyme. Dispense 18 µL per tube, then add 2 µL of sample.

These are the values the calculator opens with, so you can check its output against this example.

Assumptions

  • Each stock and its final concentration are entered in the same unit.
  • Volumes add on mixing.
  • The sample is added to each tube and is not part of the master mix.

Common mistakes

  • Preparing exactly enough for the number of reactions. Some volume is always lost, and the last tube comes up short.
  • Including the sample volume in the master mix, or forgetting to subtract it from the water.
  • Leaving enzymes out of the mix to add them tube by tube, which brings back the pipetting error the mix was meant to remove.

Related tools

PCR Master Mix CalculatorComponent volumes for any number of reactionsOpen calculatorDilution Calculator (C1V1 = C2V2)Stock and diluent volumes for any dilutionOpen calculatorReagent and Buffer Recipe CalculatorWeigh-out masses and stock volumes for a whole recipeOpen calculatorDNA Dilution CalculatorDilute a DNA or RNA stock to a working concentrationOpen calculator

Related equipment

Service documentation, failure modes and parts for the instruments this calculation is used with.