Greetings,
We are planning to extract microRNAs from rat hippocampal tissue for quantification by real time PCR, we are using Qiagen miRNeasy extraction kits and the Taqman assays.
I was advised that it is best to extract RNA from fresh samples, however due to the large sample size and the strict time frame of the experiment, I might need to store some of my samples for 10 days before working on them.
I have access to RNAlater , liquid nitrogen, and -20 and -80 freezers.
What would be the best option for sample storage?
Also, would it be best to store extracted RNA or cDNA rather than whole tissue?
Thank you.
Hello,
If I were you, I would extract RNA using Qiagen kit, then stored Them @ -80°C. When you will get all your RNAs proceed to one RT reaction then qPCR using taqman probes.
I would recommend to use LNA assays from exiqon rather than taqman assays.
Best.
Arnaud
Greetings,
I'd rather preserve tissue samples using RNAlater and -80 °C, or even -20 °C, until further processing, it works like a charm. Once your RNA is isolated, run RT to synthesize the corresponding cDNA. Store the remaining RNA at -80 °C and use your cDNA as much as you can. My recommendation to you is to keep stock RNA under ultrafreezing conditions and don't take it out until you have run out of cDNA, that way you'll minimize any possibility of damaging RNA due to excess of freeze - thawing cycles. ALSO, one vital step to protect it is using RNase inhibitor like RNaseZap or chlorine to clean your work area, and keep sample tubes on ice the whole time they are taken out of ultrafreezer.
Best regards.
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