RNA Ladder for Northern Blots

[H. Yim]

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
CH-32
4800 Sand Point Way NE
Seattle, WA 98105.