Does anyone can explain why I need to use A 260/280 to measure RNA amount? What this means?
The ratio of the absorbance at 260 and 280 nm (A260/280) is used to assess the purity of nucleic acids. For pure DNA, A260/280 is widely considered ~1.8 but has been argued to translate - due to numeric errors in the original Warburg paper - into a mix of 60% protein and 40% DNA. The ratio for pure RNA A260/280 is ~2.0. These ratios are commonly used to assess the amount of protein contamination that is left from the nucleic acid isolation process since proteins absorb at 280 nm.
The ratio of absorbance at 260 nm vs 280 nm is commonly used to assess DNA contamination of protein solutions, since proteins (in particular, the aromatic amino acids) absorb light at 280 nm. The reverse, however, is not true — it takes a relatively large amount of protein contamination to significantly affect the 260:280 ratio in a nucleic acid solution
Actually, you use the 260/280 ratio to assess the purity of nucleic acid samples, not to quantitate them (for that, OD260 is used directly).
Beware that the pH of the sample will significantly influence 260/280 measurements; hence, it is preferable to dissolve the sample in TE buffer or something similar for this purpose. See e.g. https://www.ncbi.nlm.nih.gov/pubmed?orig_db=pubmed&term=9067025
Hope it helps!
The ratio of the absorbance at 260 and 280 nm (A260/280) is used to assess the purity of nucleic acids. For pure DNA, A260/280 is widely considered ~1.8 but has been argued to translate - due to numeric errors in the original Warburg paper - into a mix of 60% protein and 40% DNA. The ratio for pure RNA A260/280 is ~2.0. These ratios are commonly used to assess the amount of protein contamination that is left from the nucleic acid isolation process since proteins absorb at 280 nm.
The ratio of absorbance at 260 nm vs 280 nm is commonly used to assess DNA contamination of protein solutions, since proteins (in particular, the aromatic amino acids) absorb light at 280 nm. The reverse, however, is not true — it takes a relatively large amount of protein contamination to significantly affect the 260:280 ratio in a nucleic acid solution
The ratio of absorbance at 260 nm and 280 nm is used to assess the purity of DNA and RNA. A ratio of ~1.8 is generally accepted as “pure” for DNA; a ratio of ~2.0 is generally accepted as “pure” for RNA.
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