On Wed, 1 Nov 1995, Hoi Pang Low wrote:
> I have very bad luck with RNA ladders in doing a Northern Blot from
> formaldehyde gels. I would appreciate hearing from anyone who can
> recommend a good RNA ladder,from which company, protocol to follow, etc.
>> Thank you very much.
I too had bad luck with RNA ladders until I started staining the ladder
AFTER blotting onto the Nylon. Run your ladder normally in the
formaldehyde gel (I use 0.66M in 1X MOPS), rinse gel in dH2O a few times
and transfer with 20X SSC. After blotting, rinse blot in 5X SSC, dry, UV
cross-link and cut off the RNA ladder lanes. Stain in 0.5M sodium
acetate (pH 5.2) for a few minutes and rinse off excess stain in dH2O.
Keep wet and wrap in saran. Its a good idea to make a photocopy of the
stained lane or mark the bands with a pen since the stained lanes will
fade. I between 2- 3 ug of BRL's RNA ladder and the only time it didn't
work was when I did an alkaline transfer. Good Luck,
Harry Yim, Ph.D.
Children's Hospital and Medical Center
Department of Infectious Diseases
4800 Sand Point Way NE
Seattle, WA 98105.