I am looking for the quick and reliable method for RNA extraction from the whole cartilage or small pre-cut discs (about 3 mm). I found many different methods in the literature. Pulverization of shock-frozen samples at the beginning sounds like a good idea, but it doesn't look as a high throughput method since you need to re-use metal vessels. I am also uncertain about which RNA isolation kit/method to use, as well as about RNA late and proteinase K treatments. It will be great to get an input from people who have a hands-on experience with cartilage.
I did some trizol extractions using that kind of method on fish vertebra (bigger sample than yours). I just wrap the sample in a little piece of foil, put it in liquid nitrogen for a few minutes and then pulverize it with a hammer. unwrap the aluminum and put the produce dust in trizol. I do a 20-30 sec of polytron, on ice, but if your sample heat, it degrade fast (in my case)
We use Qiagen's miRNEasy according to protocol, it gives us consistently high yield. Briefly, snap freeze sample in liquid nitrogen, pulverize with mortar pestle, use Qiazol then RNEasy affinity column.
Dominik, thanks for the answer. What type of mortar pestle do you use? I never pulverised tissue samples before. Do you need to clean mortar between the samples or I can find disposable?
Thank you,
I use cheap ceramic ones from Fisher, (S337621, about $65 for a set of 10), and keep the sample under liquid nitrogen while pulverizing it. I wipe everything between samples, how carefully depends on how separate I need to keep the RNA.
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