To estimate the nutritional condition of fish and invertebrate larvae, biochemical methods are widely used. One of the most important is the RNA/DNA ratio. Since the quantity of DNA remains constant in the cells regardless the condition of the individual, but RNA levels are associated to metabolic activity, this ratio provides an accurate indication of the nutritional status of the individual.
Samples collected for this purpose are generally frozen or kept in RNAlater, which avoids any RNAse activity. However, often samples are collected with other purposes and kept, for example, in ethanol. Would it be feasible to use these samples for this type of analysis?
You can extract DNA/RNA from ethanol stored tissues. Trizol method is best for RNA/DNA extraction . Long storage of samples in ethanol can effect the results better use fresh samples.
Hi David,
I am afraid that 70% ethanol might not be sufficient to inhibit RNA degradation. The two methods that are commonly used in my lab are snap freeze in liquid nitrogen followed by storage in a -80 freezer or sampling a little piece of tissue immediately after euthanasia then store in RNAlater.
Maybe this paper will help you as it provides a discussion on how good ethanol is at preserving nucleic acids.
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1850907/
good luck with your project!
Tullio
The answer is no. RNA (and even DNA) stored in water solutions progressively degrades when kept at room temperatures. Samples kept in 70% ethanol have enough water content to produce RNA degradation. In order to quantify RNA you should have kept the sample in RNAlater or frozen at -80ºC. Sorry!
Gracias Owen ;)
So, if I understand well, the main problem in this case (70% ethanol) would be with RNA, that degrades faster than DNA, isn't?
If both RNA & DNA degrade at the same rate I guess that I still could estimate the RNA/DNA ratio from each sample, which is what really interests me; I don't need to know the actual quantities of nucleic acids in the samples.
Yep, that would be your main problem. An additional problem is that RNA-degradation and DNA-degradation kinetics are not well understood yet. The fact is that RNA degrades faster than DNA, but nobody knows how faster. Many molecules and ions commonly found in biological samples may act as catalyzers or inhibitors; e.g., see http://www.sciencedirect.com/science/article/pii/0005278772905096 on quantifying the degrading action of H2O2 in the presence of several ions.
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