If I extract RNA from a (leaf tissue) sample using a one-step phenol:chloroform extraction, how long can those samples be stored at -80°C? And how many times can I defrost and refreeze them before they will become degraded?
I've found that extracted RNA using commercial kits has stayed stable for many years at -80 C. I would certainly aliquot it before freezing however as RNA is particularly sensitive to freeze-thaw cleavage.
We can keep extracted RNA in -80°C for a few weeks, but before the start of any experiments, it needs to be validated by gel electrophoresis.
I keep my RNA in 1mM sodium citrate pH 6.4. Citrate is a chelator, and helps trap the divalent metals many RNAses need to work. The lower pH also helps inhibit RNAse activity. EDTA could work as a chelator, but it only chelates well at pH's where RNAse activity is worse. Citrate gives both chelation and low pH.
Of course I also keep my RNA at -80.