Why does changinf buffer effect isolation of RNA? - (Sep/29/2006 )
Hello! (first time user..bit nervous ),
I was wondering if anyone would be able to explain a problem I am having.
I am isolating the nucleic acids from soil (in particular the RNA) using a standard bead beating with phenol method. The published protocol uses K phophate buffer. This works well for some of our soils (normal grassland bog standard soils). However one of our soils which has had NPK added to it as a fertlizer regime for over 100 years is causing a problem. I can isolate good high Mw DNA but no RNA at all. Yet when I switch the buffer from K to Na phosphate, I am able to isolate the RNA. I will hold my hands up here and admit chemistry is not my strongest subject but my understanding was that the buffer was just there to stablize the pH. Clearly it is the K or the Na which is having an effect on the isolation of the RNA. Would anyone know why this would be happening?
thanks in advance.
One of the big differences in protocols of K vs. Na is the solubility of K.SDS vs. Na.SDS salts. If your protocol uses SDS to solubilize material, excess K may precipitate the SDS before it can do its job, whereas the sodlum has little effect. Just a wild guess.