For study with RNA, we made two samples one with annealed at 90 degree for 1 min and other one is non-annealed. Now, we found the annealed one didn't give us the expected result whereas the non-annealed one gave the expected result. So, I want to know what structural changes occur in the RNA when it is heated or annealed ?
My experience is with purification of RNA using dHPLC. At 90’C the RNA will melt to become single stranded. The quality of double stranded product on re-annealing has to do with rate of cooling, the fragment lengths, and the competing sequences that may be present competing for strand association, if this is what you mean.
David Hild Thank you for sharing your opinion. We are trying to attach a biotynilated mRNA to a Streptavidin surface; on our surprise we got a nice attachment with the non-annealed / non-heated mRNA, but with the annealed one we didn;t get any binding at all. Our possible explanation goes like may be somehow after annealing the mRNA aquries a structure where the biotynilated end is no more accessible to the Streptavidin surface to bind with; or somehow we got contaminated with RNAse. What is your opinion regarding this aspect ?
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