My hypothesis: Since nucleic acids has a peak absorbance at 260nm wavelength and proteins at 280nm, the ratio of greater than 2.0 is considered relatively pure RNA.
Twist: However, literature indicates that 260/280 ratio for relatively pure RNA is approximately 2.0. (What about having 2.4 or 2.5?)
Other concern: could a > or = to 2.0 also indicate purity from DNA contamination?
For RNA, maximal absorbance is measured at 260 nm. At 280 nm, RNA absorbance is 2 fold less. This explain that 260/280 ratio should be 2 for pure RNA. In case of protein contamination, A280 increase then the ratio 260/280 decrease.
I'm not an expert but I guess that ratio > 2 can be due to contaminant absorbing at 260 nm... for example CTAB (component of some lysis buffers). Moreover, according to Nandrop manual, pH can also influence 260/280 ratio. Acidic solutions will under-represent 260/280 ratio (by 0.2 to 0.3), and basic solution will over-represent the ratio.
Values of ~ 2.0 is generally accepted as pure for RNA.
See those links, that might help you:
http://biomedicalgenomics.org/RNA_quality_control.html
http://www.bio.davidson.edu/projects/gcat/protocols/NanoDrop_tip.pdf
For RNA, maximal absorbance is measured at 260 nm. At 280 nm, RNA absorbance is 2 fold less. This explain that 260/280 ratio should be 2 for pure RNA. In case of protein contamination, A280 increase then the ratio 260/280 decrease.
I'm not an expert but I guess that ratio > 2 can be due to contaminant absorbing at 260 nm... for example CTAB (component of some lysis buffers). Moreover, according to Nandrop manual, pH can also influence 260/280 ratio. Acidic solutions will under-represent 260/280 ratio (by 0.2 to 0.3), and basic solution will over-represent the ratio.
Bear in mind that the 260/280 ratio is not a simple measure of NA purity. Its purpose was to detect the contamination of protein with NA, for which it is very sensitive. It is not at all sensitive to the contamination of NA with protein since it takes a relatively large amount of protein contamination to significantly affect the 260:280 ratio in a nucleic acid solution.
http://www.biotek.com/resources/docs/Measure_Your_Purity_GIT_MayJune2008.pdf
- Badges
- Science topic
- Similar topics
- Biological Science
- Molecular Biology
- Biomolecules
- Nucleic Acids