DNA Concentration Calculator
Calculate nucleic acid concentration from spectrophotometry (A260) readings using the Beer-Lambert Law.
Sample & Instrument Settings
Formula Used (Beer-Lambert Law)
Understanding DNA Concentration Measurement
Accurate DNA quantification is a cornerstone of molecular biology. Whether you’re preparing samples for PCR, Next-Generation Sequencing (NGS), cloning, or transfection, knowing your exact nucleic acid concentration is critical for reproducible results.
How Does This Calculator Work?
This calculator uses the Beer-Lambert Law, the gold standard equation in spectrophotometry:
C = (A260 × Conversion Factor × Dilution Factor) / Pathlength
Where C is the concentration in μg/mL.
When UV light at 260 nm passes through a solution containing nucleic acids, the aromatic bases absorb that light. The degree of absorbance is directly proportional to the concentration of the nucleic acid present.
Standard Conversion Factors
- Double-stranded DNA (dsDNA): 50 μg/mL per A260 unit — the most common for genomic DNA.
- Single-stranded DNA (ssDNA): 33 μg/mL per A260 unit — used for plasmids or single-stranded oligos.
- RNA: 40 μg/mL per A260 unit — the slightly different factor accounts for the unique stacking interactions of RNA bases.
- Custom Oligonucleotides: The factor must be calculated from the specific sequence composition using the nearest-neighbor model.
Purity Ratios Explained
Spectrophotometry can also give insight into sample purity by comparing absorbances at different wavelengths.
- A260/A280 Ratio: Proteins absorb strongly at 280 nm. A ratio of ~1.8 for DNA and ~2.0 for RNA indicates a pure sample. Values significantly below these suggest protein contamination.
- A260/A230 Ratio: Values around 2.0–2.2 are ideal. Low ratios indicate contamination from organic solvents (phenol, guanidinium), EDTA, or salts commonly found in extraction buffers.
Common Sources of Error
- Absorbance out of range: Readings below 0.1 or above 1.0 are unreliable. Dilute or concentrate your sample accordingly.
- Incorrect blank/reference: Always blank your instrument with the same buffer used to dissolve your nucleic acid, not just water.
- Wrong sample type selected: Using the dsDNA factor for RNA will give a 20% error in your concentration.
Data Sources & References
- Beer-Lambert Law and A260 DNA quantification: Sambrook, J. & Russell, D.W. (2001). Molecular Cloning: A Laboratory Manual, 3rd ed. Cold Spring Harbor Laboratory Press. Appendix 8.
- Extinction coefficients for dsDNA (ε = 50 ng·cm/μL), ssDNA (ε = 33), and RNA (ε = 40): Thermo Fisher Scientific Technical Reference. thermofisher.com
- Avogadro constant (6.02214076×10²³ mol⁻¹): CODATA 2018, NIST SP 961. physics.nist.gov
- Average molecular weight of base pair (650 g/mol): Sambrook & Russell (2001), Table A8.1.
