How It Works
This calculator works in 4 steps.
List the oligos
Type forward, reverse, and probe names with stock concentrations.
Enter the target per oligo
Type the concentration each oligo should have in the mix.
Enter the mix volume
Type the total mix volume.
Read the volumes
The calculator returns each oligo volume, the water volume, and the per-oligo concentration.
The Formula
The formula applies C₁V₁ = C₂V₂ to each oligo against the full mix volume.
Vᵢ = Cᵢ,target × V_mix / Cᵢ,stockV_water = V_mix − Σ VᵢC_rxn = C_mix × V_added / V_rxn- Cᵢ,stock:
- Stock of oligo i
- Cᵢ,target:
- Concentration of oligo i in the mix
- V_mix:
- Mix volume
A 200 µL mix at 5 µM each from 100 µM stocks needs 10 µL forward, 10 µL reverse, and 180 µL water.
Conversion Table
The conversion table gives oligo and water volumes for 5 forward-reverse primer mixes.
| Stocks | Target in mix | Mix volume | Primer volumes | Water |
|---|---|---|---|---|
| 100 µM each | 10 µM each | 100 µL | 10 µL F + 10 µL R | 80 µL |
| 100 µM each | 5 µM each | 200 µL | 10 µL F + 10 µL R | 180 µL |
| 100 µM each | 20 µM each | 100 µL | 20 µL F + 20 µL R | 60 µL |
| 100 µM each | 2 µM each | 500 µL | 10 µL F + 10 µL R | 480 µL |
| 50 µM each | 10 µM each | 100 µL | 20 µL F + 20 µL R | 60 µL |
What Is the Primer Mix Calculator?
The Primer Mix Calculator is a tool that combines forward and reverse primers, and optional probes, into one premixed stock and returns the concentration of each oligo in the mix. Each oligo is diluted by the full mix volume, not by the volume of the other oligos alone.
A primer mix halves pipetting steps in every reaction. Mixing 10 µL of 100 µM forward, 10 µL of 100 µM reverse, and 80 µL water gives 10 µM of each primer, not 20 µM combined. The calculator lists each oligo's concentration on its own line to prevent that mistake.
Common Uses
There are 5 main uses of this calculator.
qPCR assays
Labs combine primers and probe into a 20× assay mix.
Genotyping panels
Users prepare primer pairs for many loci.
Multiplex PCR
Researchers balance several primer pairs in one mix.
Teaching kits
Instructors prepare premixed primers for student groups.
High-throughput screens
Core labs reduce dispensing steps with premixes.