I need to know the best way to preserve RNA extract from tissue for a long period.
Well, DEPC (treated) water and nuclease free water are basically the same thing.
DEPC water is when you "treat" your water with DEPC to denature RNases, then you autoclave the water to degrade the DEPC. You want to degrade it because DEPC can degrade nucleic acid and inhibit RT reactions (denatures the enzyme).
Nuclease water is basically water that you buy that has no RNases or DNases. Different brands use different methods I think but all give you the same water with no nucleases.
Therefore you can use any of them, just make sure you aliquot your stocks so you do not contamine them with nucleases (or bacteria that will generate nucleases) by repeteaded uses.
Hi,
RNA could be damaged even at -20 Celsius. For long term storage, I'll recommend preserving RNA at lower than -70 Celsius.
Good luck!
https://www.thermofisher.com/tr/en/home/brands/product-brand/rnalater.html
Stored tissue in RNALater for several years, was fine for both RNA and protein. Also do recommend a deep freezer as Hao pointed out.
Hi
there are two ways that you can store your sample.
1. Collect the tissue samples in liquid nitrogen later store them in -80'c.
2. You can also isolate the RNA from tissue and store @-80'c in DEPC water, ethanol, or even formamide. If you are dissolving in DEPC water, you can also add RNAse inhibitor which porotects from residual RNAse contamination.
Also remember to keep isolated RNA samples in aliquots so you can avoid major degradation by thawing and freezing cycle.
Hope this will help you!
I already stored the tissue in RNA later. But i was asking about after the extraction before i perform the RT-step. Its better to work the RT of all samples at once, right? That's why i wanted to know the best way to preserve the extracted RNA. Does DEPC water really preserve THE RNA better than nuclease free water? In the elution step should i replace the nuclease free water with DEPC?
Well, DEPC (treated) water and nuclease free water are basically the same thing.
DEPC water is when you "treat" your water with DEPC to denature RNases, then you autoclave the water to degrade the DEPC. You want to degrade it because DEPC can degrade nucleic acid and inhibit RT reactions (denatures the enzyme).
Nuclease water is basically water that you buy that has no RNases or DNases. Different brands use different methods I think but all give you the same water with no nucleases.
Therefore you can use any of them, just make sure you aliquot your stocks so you do not contamine them with nucleases (or bacteria that will generate nucleases) by repeteaded uses.
How long can i store the RNA after extraction in -80 without affecting the yielded RNA?
Hi
You can elute either in DEPC or Nuclease free water will be the same.
While I was working with samples collected in RNA later I used too add RNase inhibitor into the RNA collecting vial before elution .
If you have doubts about using RNAse Inhibitor, you can check out this discussion link.
https://www.researchgate.net/post/Does_adding_RNase_inhibitor_to_freshly_extracted_RNA_in_RNAse-free_water_inhibit_subsequent_RT_reactions
About Storing
I have an experience working with RNA stored for a year and still have better results. But keep in mind that you must avoid too many times thawing and freezing. So better maintain in aliquots.
Remember if you can do RT it will be more easy and secure to store cDNA for a long time even in -20'c than RNA always.
- Badges
- Science topic
- Similar topics
- Phytochemistry
- Extracts