PCR Master Mix Calculator
Scale PCR master mix, primers and water across reactions with overage, using C1V1=C2V2 for primer volume and keeping template separate.
Calculate PCR master mix volumes
Scale common PCR components across reactions with an overage allowance and concentration-based primer calculation.
| Component | Per reaction | Batch volume |
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How the PCR Master Mix Calculator works
The calculator derives each primer volume from stock concentration × volume = final concentration × final reaction volume. It then subtracts 2× master mix, both primer volumes and template from total reaction volume to calculate water. The shared cocktail is scaled by reaction count plus the entered overage; template is shown separately.
How to use this pcr master mix calculator
Enter your validated assay volumes and concentrations rather than treating the defaults as a protocol. Thermo Fisher PCR guidance shows the same C1V1=C2V2 approach for primer/probe calculation and often scales non-template components as a master mix to reduce pipetting variability. Choose an overage suitable for your plate size and pipetting losses.
How to interpret the result
The batch-reaction equivalent makes overage transparent. For 24 reactions with 10% overage, common cocktail components are prepared for 26.4 reaction equivalents. Template volume is not multiplied by overage in the shared cocktail because each sample/standard is typically added to its own reaction.
Assumptions and limitations
PCR formulations differ by polymerase mix, primer concentration, probes, MgCl2, additives and sample volume. Some kits have minimum/maximum template fractions. This calculator includes only two primers, 2× master mix, water and template; adapt your laboratory worksheet when additional components are required and always follow validated kit/SOP instructions.
Practical example and workflow
For a 20 µL reaction using 10 µL of 2× mix, 10 µM primers at 0.5 µM final, and 2 µL template, each primer contributes 1 µL and water is 6 µL. A 24-reaction run with 10% overage scales the shared 18 µL cocktail to 26.4 reaction equivalents.