I am using a kit for RNA extraction from Promega. We have good results for leaf, but with root the isolation is not good, 6ug/mL. May soil residues interfere with extraction? How many grams of tissue do you use?
Hi Edgardo
What you can try to increase the yields of your extractions is to add a bit of Polyclar (a Polyvinylpolypyrrolidone to bind phenols which interfere with the extraction) to the roots before you grind them. Since we are using Polyclar for our extraction we obtained much better yields. We are using lyophilized roots for extractions of around 20-50mg, if you are using frozen roots, you should not use more than 100mg.
Dear, thanks for your reply. Can you tell me which method do you use? I am using TRIZOL Reagent. It has phenol, so i dont sure that if i can use PVPP.
Dear Edgardo
It might interfere, but as you add the PVPP and grind the roots very well before you add the extraction buffer containing the TRIZOL, it should work anyway. Maybe you should pay attention that you don't put to much of the PVPP, you could try some different amounts to find the best rate.
So far, I used mainly the RNeasy plant mini kit of Quiagen which worked best for roots grown in soil. Once I also tried the Trizol method but as the yield were not very high, I sticked to the kits for RNA extraction.
Recently I tried a method to extract DNA and RNA simultaneously from soil without kit and it worked well. It was the DNA/RNA Coextraction method after Griffiths and Whiteley et al., 2000 using CTAP, would be another option if you would have access to CTAP and the TRIZOL method with PVPP would fail.
Hope you will manage to overcome the problems. Best wishes.
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