RNA sample stuck in well during agarose electrophoresis - (Nov/21/2005 )
Today I ran a 1.5% agarose gel in 1X TBE stained with EtBr. The ladder ran out fine, but the RNA samples seem to have stayed in the well. I got a relatively strong signal from the wells, but no where else. Unfortunately, I can't download the picture. Any ideas??
The RNA is from formalin fixed, paraffin embedded tissues, and was extracted following Ambion's Optimum Kit. I am concerned with the precipitate in the DNase inactivating buffer that is the last step of the kit, I couldn't get the pellet to stay and probably took some of the precipitate along with the eluted RNA.. could this be making it stick in the well? Can I filter the RNA through a spin column to get rid of the precip, or will this be bad for my RNA??
Thanks!
-jwahini-
QUOTE (jwahini @ Nov 21 2005, 04:43 PM)
Today I ran a 1.5% agarose gel in 1X TBE stained with EtBr. The ladder ran out fine, but the RNA samples seem to have stayed in the well. I got a relatively strong signal from the wells, but no where else. Unfortunately, I can't download the picture. Any ideas??
The RNA is from formalin fixed, paraffin embedded tissues, and was extracted following Ambion's Optimum Kit. I am concerned with the precipitate in the DNase inactivating buffer that is the last step of the kit, I couldn't get the pellet to stay and probably took some of the precipitate along with the eluted RNA.. could this be making it stick in the well? Can I filter the RNA through a spin column to get rid of the precip, or will this be bad for my RNA??
Thanks!
The RNA is from formalin fixed, paraffin embedded tissues, and was extracted following Ambion's Optimum Kit. I am concerned with the precipitate in the DNase inactivating buffer that is the last step of the kit, I couldn't get the pellet to stay and probably took some of the precipitate along with the eluted RNA.. could this be making it stick in the well? Can I filter the RNA through a spin column to get rid of the precip, or will this be bad for my RNA??
Thanks!
Do you put your eluted RNA with loading buffer on ice for too long after boiling it at 65 degree? I find out that if I put the RNA on ice for too long, the RNA precipate. After I load onto the gel, the RNA stuck in the well.
I hope this may help.
-Minnie Mouse-