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MasterMath

DNA Concentration Calculator

The concentration of DNA or RNA from the absorbance at 260 nm, with the 260/280 ratio that tells you whether the sample carries protein. The factor depends on the nucleic acid and is chosen in the form.

Concentration

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Concentration—
260/280 ratio—
Purity—
Factor applied—
In 50 µl of sample—

How this was worked out

    The formula

    C = A₂₆₀ × factor × dilution

    What it means

    Nucleic acids absorb most strongly at 260 nanometres, and in the range where the Beer-Lambert law holds, absorbance is proportional to concentration. The constant of proportionality depends on what the sample is: an absorbance of 1 corresponds to about 50 ng/µl of double-stranded DNA, 40 of RNA and 33 of a single-stranded oligonucleotide. Proteins absorb at 280, so the ratio between the two readings measures how clean the preparation is.

    How to work it out by hand

    1. Measure the absorbance at 260 nm, blanked against the same buffer
    2. Multiply by the factor for the nucleic acid you are measuring
    3. Multiply by the dilution if you measured a diluted sample
    4. Divide the 260 reading by the 280 one to check purity

    What is worth knowing

    The measurement is only reliable with an absorbance at 260 between 0.1 and 1: below that noise takes over and above it the detector saturates, and in both cases you have to dilute and measure again. A 260/280 ratio around 1.8 is good for DNA and 2.0 for RNA; below 1.7 there is protein or phenol. And this measures everything that absorbs at 260, not only what you care about: free nucleotides and contaminating RNA count the same, so a high concentration does not mean the DNA is intact.

    Frequently asked questions

    Which factor should I use?

    50 for double-stranded DNA, 40 for RNA and 33 for single-stranded oligonucleotides. You pick it at the top of the form.

    What 260/280 ratio is good?

    Around 1.8 for DNA and 2.0 for RNA. Below 1.7 there is usually protein or leftover phenol.

    What if the absorbance is above 1?

    Dilute the sample and measure again. Above 1 the detector stops being linear and the reading is not usable.

    Does it measure only my DNA?

    No: it measures everything that absorbs at 260 nm. Free nucleotides and contaminating RNA add to it, so the figure can sit above the real one.

    Why is the concentration in ng/µl?

    That is the unit microvolume spectrophotometers work in. One ng/µl is the same as one µg/ml.