Sorry if this is a basic question, but it is something that I was unsure about. I am extracting long hairpin dsRNA from HT115 E. coli for some RNAi experiments. The ssRNA from the expression cassette should form a long hairpin dsRNA structure. However, I think that there are some mismatched bases in the stem loop. Would this mean that the stem loop could potentially be cleaved by ssRNA-specific endonuclease? I appreciate your help and apologies if it's a basic question. Nonetheless, thanks for sharing your wisdom.
Hi Antonio, I have observed RNaseA cleave dsRNA in the past, but only under low-salt conditions (below 100mM I believe). I assume it would be the same for RNase T1 because they function very similarly. In my experiments, I used short oligos (27-60bp) so this might vary for you. Overall, I would estimate that both RNAseA and T1 will cleave the stem under low salt conditions. My oligos were usually completely digested when using low salt. Here is a publication that might help Article Degradation of Double-Stranded RNA by Mammalian Pancreatic-T...
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