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Star Republic: Guide for Biologists

Detection of DNA/RNA --- spectrometer

1. Turn on OD absorbance spectrometer. Turn on UV source. Wait 5-10 minutes for the UV source to stabilize.

2. Rinse the quartz measurement cuvette with dH2O. Note plastic cuvettes are not UV transparent, thus can't be used for UV measurement.

3. Adjust UV wavelength to 260 nm, fill the quartz cuvette with water. Zero the spectrometer, i.e., the OD reading for water should be 0.

4. Measure UV absorbance at 260 nm for the sample.

5. To correct for protein contaimnations, repeat steps 2-4 at the wavelength 280.

6. Calculate DNA/RNA concentration using the OD calculator of BioToolKit or use the following estimations (for cuvette with 1-cm light path):

  • Double-stranded DNA concentration (mg/ml) = 50 * A260
  • Single-stranded DNA/RNA concentration (mg/ml) = 40 * A260
  • Oligonucleotides concentration (mg/ml) = 33 * A260
  • 7. A260/A280 should be within 1.8-2.0. Otherwise the protein contamination is too high.

    8. Clean up the cuvette, turn off the UV source and the spectrometer.