Answer to Question #200749 in Biochemistry for pop

Question

• 2) Why are the ratios of OD260/OD280 and OD260/OD280 for clean nucleic acids about 2.0? Show your answer by drawing and explaining a DNA absorbance spectrum from 200 to 300 nm. Which type of contaminations can you detect with these measurements? 

Answer 1

If the ratio of DNA OD260/OD280 is between 1.8 and 2.0, the DNA purity (free from protein contaminants) is ~90% or better. If the ratio of RNA OD260/OD280 is ~2.0, the RNA purity is ~90% or better.

260/230

This ratio is used as a secondary measure of nucleic acid purity. The 260/230 values for “pure” nucleic acid are often higher than the respective 260/280 values. Expected 260/230 values are commonly in the range of 2.0-2.2. If the ratio is appreciably lower than expected, it may indicate the presence of contaminants which absorb at 230 nm.

The most common purity calculation is the ratio of the absorbance at 260nm divided by the reading at 280nm. Good-quality DNA will have an A₂₆₀/A₂₈₀ ratio of 1.8–2.0.