If the RNA is complete in solution, the concentration should be the same.
But, if you extract the RNA and resolved the pellet not completly (which could sometimes happen), the concentration could be higher after thawing. You can use freezing/thawing to resuspend RNA pellet.
Dear Abdelraheim,
It is quite well established that nucleic acids can get stuck to plastic ware such as microcentrifuge tubes unspecifically. This is usually not noticeable at high conc., but can be significant at lower conc. You can reduce this effect by adding glycogen to low conc. RNA (if this will not interfere with downstream applications) and also by using specific low binding tubes.
Thank you Christian Tüting,
and thank you Petra Disterer , but I did not hear about adding glycogen to low conc. RNA and I think I may use it. Could you please guide me adding it?
Abdelraheim,
Glycogen is usually added to low conc. RNA to aid precipitation https://www.thermofisher.com/order/catalog/product/R0551
but it also prevents the RNA from binding unspecifically to the plastic as the RNA will be attached to the glycogen.
I strongly advise against using freeze/thaw cycles to aid resuspension of RNA pellets. This may be fine if you are absolutely sure that there are no RNases in your solution, but that is likely not the case.
In fact, this is an alternative explanation for loss of RNA - you simply have some RNase contamination that "ate" your RNA. However, if you measure the conc. by Nanodrop you would still see the original conc. as the nucleotides wouldn't simply evaporate into thin air but remain in the solution (Nanodrop measures conc. of nucleotides NOT RNA or DNA! So a Nanodrop measurement of 50 ng/ul could be 40 ng/ul of nucleotides and 10 ng/ul of an actual RNA/DNA sequence).
For the resuspension of RNA you should use DEPC treated ultrapure water. The DEPC will inactivate the RNase and you will have no degradation.
To check if your sample is degraded, you can simply analyse via agarose-gel.
It's explained here: http://bitesizebio.com/13527/how-to-quality-control-check-your-rna-samples/
you have to check the rRNA bands.
I personally never use DEPC treated water with RNA as it can inhibit the RT step. RNAse free water is my choice.
Also, DEPC is carcinogenic (it carboxymethylates purines) and should be handled with care. Wear gloves! If you need to then Di-methyl-propyl carbonate (DMPC) is a safer alternative to DEPC. It's is used in exactly the same way.
https://www.thermofisher.com/uk/en/home/references/ambion-tech-support/nuclease-enzymes/tech-notes/rnase-and-depc-treatment.html
http://www.thermofisher.com/uk/en/home/references/ambion-tech-support/nuclease-enzymes/general-articles/the-basics-rnase-control.html
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