Hi all I have a question about RNase A and column vs non column minpreps.
In Qiagen RNase A is added to the initial resuspension buffer. I am unsure
as to what good this does as in P1 the RNA/DNA is still in the bacterial
cells. When the alkaline lysis buffer is added in P2 the DNA/RNA is
released but I read RNase A is inactivated in .1% SDS (P2 is 1% SDS). In
any case RNA generally does not bind to the silica columns that much anyways
so the columns remove the RNA.
I have been using the Qiagen buffer to do the alkaline lysis then doing
phenol/chloroform extraction followed by ethanol precip. When I do this at
the end there is a bright RNA band on the gel around 50-100bp. Qiagen
claims in their manual this is RNase A-resistant RNA.
My question is will precipitating the DNA/RNA after the lysis and
resuspending in TE/RNase then doing PC extraction and another alcohol
precipitation remove the RNA because RNase A is inactivated in SDS or is
Qiagen correct in saying this RNA is simply resistant?