What it does
A spectrophotometer gives absorbance; you need concentration, a purity check and, for PCR, cloning or sequencing, the molarity or copy number and how to dilute. This page does all of it for one sample or a whole batch, in your browser.
- Concentration = (A260 − A320) × factor × dilution ÷ path length, in ng/µL. The factor is 50 for double-stranded DNA, 33 for single-stranded DNA and 40 for RNA; you can change it (short oligos need a sequence-specific factor from the supplier).
- Protein uses A280 with your extinction coefficient ε and molar mass: c = A × M ÷ (ε × path), in mg/mL and µM.
- Purity: the 260/280 and 260/230 ratios, background-corrected, with a plain note when they fall outside the usual ranges, and a chart of both ratios.
- Molarity and copies from the length (average base mass) or the exact sequence, for linear or circular DNA.
- Dilution: stock and diluent volumes to reach a target in ng/µL, nM or copies/µL (mg/mL or µM for protein).
How to use it
- Sample type. Choose double-stranded DNA, single-stranded DNA, RNA or protein. Set the dilution if you diluted before measuring, and the path length (leave 1 cm for microvolume readers, which already scale to 1 cm).
- Readings. One row per sample: name, A260, A280, then A230 and A320 if you have them. Paste, type or open a CSV or spreadsheet.
- Size. Give the length in bp or nt, and the shape for double-stranded DNA. Paste the sequence instead for its exact mass.
- Dilute to. Pick a target, a unit and the final volume.
Worked example
The made-up example has four double-stranded DNA preps of a 3,000 bp linear fragment. Prep 1 reads A260 7.40 and A320 0.02, so it is (7.40 − 0.02) × 50 = 369 ng/µL. A 3,000 bp linear double strand weighs about 1,853,733 g/mol, so that is 199 nM, or 1.20 × 10¹¹ copies/µL. To make 50 µL at 10 ng/µL, mix 1.36 µL of stock with 48.6 µL of water.
Prep 3 has a 260/280 of 1.62, below the usual 1.7 for DNA, which hints at protein or phenol left over. Prep 4 has a 260/230 of 1.39, which hints at salt or guanidine carried over from the kit. The tool flags both: “4 samples, 43–369 ng/µL; 2 to check”.
Good to know
- Below about 0.02 absorbance units readings are unreliable, and the tool says so.
- A260 counts every nucleic acid, so RNA in a DNA prep (or free nucleotides) inflates the result. A dye-based assay is more specific.
- The purity ranges are rules of thumb; ratios also shift with the pH and salt of the blank.
FAQ
Why subtract A320? Nucleic acids and proteins barely absorb at 320 nm, so a reading there is background from turbidity or the optics.
Does circular DNA weigh less? Slightly: a closed circle has no free ends, so it lacks the two water molecules of a linear double strand. The difference matters only for short molecules.
Is anything uploaded? No. Everything runs in your browser.
Where the numbers come from
- Beer–Lambert law with the conventional factors of 50, 33 and 40 µg/mL per A260 unit for double-stranded DNA, single-stranded DNA and RNA (general laboratory facts). Purity ranges are common rules of thumb, not a standard.
- Nucleotide residue masses worked out by fibub from the chemical formulas with standard atomic weights; copies use the Avogadro constant (exact SI value).
- Checked in the tests against an independent Python calculation. The example readings are made up.
Standard scientific values, such as constants, atomic weights and melting points, are facts found in every textbook; we name where ours were checked. If you are an author or publisher and want a value, a credit or a table changed or removed, tell us and we will act quickly.
Last updated 2026-11-09.