DNA copy number calculator
Maintained by the Talindrew team · Last updated
The number of DNA copies in a sample is the mass divided by the per-molecule weight, times Avogadro's number: copies = ng × 6.022 × 10²³ / (length in bp × 660 × 10⁹). One nanogram of a 3,000 bp plasmid contains about 3.0 × 10⁸ copies: the arithmetic behind every qPCR standard curve.
Result
copies = ng × 6.022×10²³ / (bp × 660 × 10⁹), assuming double-stranded template at 660 g/mol per bp. 1 ng of a 3 kb plasmid ≈ 3.0 × 10⁸ copies.
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What it looks like
Formula
copies = ng × 6.022×10²³ / (N bp × 660 × 10⁹)
Source: Avogadro constant; 660 g/mol per bp (Thermo Fisher copy-number calculator convention)
Worked example
Given: 1 ng of a 3,000 bp plasmid standard
- 1.Per-molecule weight: 3,000 × 660 = 1.98 × 10⁶ g/mol.
- 2.Moles: 1 × 10⁻⁹ g / 1.98 × 10⁶ g/mol = 5.05 × 10⁻¹⁶ mol.
- 3.Copies: 5.05 × 10⁻¹⁶ × 6.022 × 10²³ = 3.04 × 10⁸.
1 ng of 3 kb plasmid ≈ 3.0 × 10⁸ copies
How the calculation flows
Units & constants
| Constant | 660 g/mol per bp (some tools use 650: ~1.5% difference; stated so results are reproducible) |
|---|---|
| Avogadro | 6.02214076 × 10²³ mol⁻¹ (exact) |
| Template | Use the FULL plasmid length, not just the amplicon |
| Typical standard curve | 10⁷ → 10¹ copies in 10-fold steps |
| Price | Free |
How do you make a qPCR standard curve from this number?
Quantify the purified standard (A260 or fluorometry), convert to copies/µl with this calculator, dilute to a convenient anchor (say 10⁸ copies/µl), then run six to eight 10-fold serial dilutions. Plot Ct against log₁₀(copies): the slope gives amplification efficiency (see the qPCR efficiency calculator) and the line converts unknowns' Ct values into absolute copy numbers.
Which length goes in: plasmid or insert?
The full length of the molecule you weighed. A 100 bp amplicon cloned in a 3 kb vector still weighs like 3.1 kb per copy, so using the amplicon length would overstate copy number thirty-fold. For linearised plasmids the length is unchanged; for a purified PCR product, use the product length.
Single-stranded templates and RNA
For ssDNA use 330 g/mol per nucleotide (half the duplex weight); for ssRNA standards use 340 g/mol per nucleotide. In vitro-transcribed RNA standards additionally need DNase treatment before quantification, or the leftover template inflates the count.
Frequently asked questions
About 9.1 × 10⁸: 1 × 10⁻⁹ / (1,000 × 660) × 6.022 × 10²³. Shorter molecules mean more copies per nanogram, inversely proportional to length.
The other tool likely uses 650 g/mol per bp instead of 660. Both are published averages; the discrepancy is far below quantification error. This page states its constant so you can reproduce either.
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