Help with EMSA minigel conditions

P.J. Shaw pjs14 at le.ac.uk
Thu Oct 16 11:35:35 EST 1997


Why not try 5/cm constant voltage? Pre-run the gel first for an hour or so
until the current drops to a constant level and load. For a mini-protean, 
I guess this would be about 40-50 V. I've found that for some
interactions, they can be destabilised (DNA stuck in top of wells) if you 
run >10V/cm, using 0.5X TGE without recirculation.

Philip.

pjs14 at le.ac.uk
 

On Thu, 16 Oct 1997,
Darren Tyson wrote:

> Howdy,
> 
> I'm trying to convert my electrophoretic mobility shift assay (EMSA)
> from a large gel format (20 cm X 20 cm) to a minigel format (BioRad
> Mini-Protean II).  For the large gel format the running conditions were
> constant 35 mA at 4 deg C for 4 hours.  I believe constant 35 mA for a
> minigel would be too much power for the protein-DNA interactions to be
> maintained and would finish the run very quickly.  Can anyone suggest
> the proper running conditions for my application?  BTW, I'm using a high
> ionic strength buffer (Tris/glycine/EDTA).
> 
> Any help would be greatly appreciated.  Thanks in advance.
> 
> Darren
> 
> 
> 




More information about the Methods mailing list