Hello everyone,
It's my first time working with RNA but I am told it's very easily degraded. A protocol for cDNA synthesis requires "nuclease free water". Should I use PCR water or molecular grade water? I have many RNA samples so can I prepare this water in the lab myself or should I buy commercial water?
Hi Gina,
For RNA samples, you can use DEPC treated autoclaved mili-Q water.
You can find DEPC treated nucleus free water. Available from many bio pharmaceutical companies...
Almost all of this "degradation" talk is pure superstition. If anyone lectures you about it, ask them when they have last tested it, using a well-designed experiment, with multiple different water sources side-by-side, under otherwise identical conditions. In practice, for a regular qPCR application, even tap water will work fine, and so will any source of reasonable lab water you have around.
Gina M Accumanno
PCR water or molecular grade water
It doesn't matter (until and unless there is some contamination in your water)
Both are supposed to be nuclease-free.
Hi Isha Pandey
Thank you for responding to my question.
Is there specific protocol to do this in the lab?
you can use NFW (Nuclease Free Water), DEPC treated water, Molecular Grade Water they all are the same.
Generally, PCR kits come up with vials of nuclease free water or you can buy commercially to stock it up. Other alternative is to use DEPC treated MQ water which is also nuclease free.
RNase is more stable and are more difficult to inactivate and it usually do not require auxiliary cofactor. A small amount of RNase is enough to destroy RNA. RNase exists in saliva, sweat and skin surface.
To make sure your experience is smooth…. Here are my suggestions.
1. Wear mask , gloves to avoid RNase contamination.
2. Use exclusive tools, for RNA operation only.
3. Use Disposable supplies ( RNase free tip, tube..).
4. Use RNase free water
.
1:( nuclease-free). you can use DEPC treated MQ water which is also nuclease-free.
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