I obtained a good concentration of RNA and a good ratio 260/280 after the measurement by nanodrop. Infortunately I found that the ratio 260/230 was still very low (around the average of 0.9)
Thanks and happy new year
The answer is very simple: Don't worry about it!
The common interpretation is that 260/230 ratio is indicative of small molecule contamination, such as salt, ethanol etc. Firstly, a ratio of 0.9 is not too bad. Secondly, I have done thousands (if not tens of thousands) of RNA extraction followed by lots of different assays, from direct labelling, through reverse transcription to ligation, and never found a high OD230 to have any detrimental effect. Even R&D departments of several companies who pride themselves on RNA products (e.g. Ambion, QIAGEN) have no idea what meaning to assign to 260/230 ratios!
Good luck, and Happy New Year :-)
The answer is very simple: Don't worry about it!
The common interpretation is that 260/230 ratio is indicative of small molecule contamination, such as salt, ethanol etc. Firstly, a ratio of 0.9 is not too bad. Secondly, I have done thousands (if not tens of thousands) of RNA extraction followed by lots of different assays, from direct labelling, through reverse transcription to ligation, and never found a high OD230 to have any detrimental effect. Even R&D departments of several companies who pride themselves on RNA products (e.g. Ambion, QIAGEN) have no idea what meaning to assign to 260/230 ratios!
Good luck, and Happy New Year :-)
Thanks, David. Good point re carbohydrates - I never think of plants *mental slap on the wrist*. My experience is with extracting RNA from human adherent and suspension cell lines, fresh-frozen solid tumours and FFPE-embedded archival tissues. I never truly investigated which component causes the elevated OD230, but there are no substantial carbohydrates in our system. More to the point, there was an obvious correlation between beginners' preps that smell of ethanol and high OD230 - probably due to a carry-over of the lysis/wash buffers after incomplete spins/drying/etc. Could e.g. guanidium, SDS or butanol be the culprit? Half the time the components are not even disclosed, so it is anyone's guess. The ethanol I add myself.
Either way, with the caveat of never having had carbohydrate overload (Christmas pudding aside!), I stand by my previous statement of having never witnessed a detrimental effect of higher OD230. David, have you ever observed any measurable inhibition of downstream applications?
And, sadly, the lack of knowledge in R&D departments is 100% factual.
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