agarose gel electrophoresis conditions

Joseph C. Bagshaw jbagshaw at wpi.edu
Sat Jan 3 08:43:31 EST 1998


Paul,

I will assume the "things" you are trying to separate are DNA fragments.
In my experience, fragments in the 8-14 kb range are no problem.  Ordinary
agarose in TBE buffer should do.  I would suggest at least 0.8% or even 1%
agarose; I've never gotten decent results with less.  Although TBE is more
expensive than TAE, it gives slightly better resolution in the mid to high
kb range and way better in the sub-2 kb range.  If you need really fine
resolution in the 8+ range the most important thing in my experience is to
run long and slow.  Run a 20 cm gel at 20-30 V for 24 hrs or more, or a
mini-gel at 10 V.  Also, if you need resolution of a complex mixture, e.g.
a genomic DNA digest, it helps to use a comb with wide, thin teeth, e.g
15X1 mm.  You should be able to see separation of fragments that differ by
5%, maybe less.

********************  HAVE GENES, WILL TRAVEL  ********************
Joe Bagshaw, Worcester Polytechnic Institute
jbagshaw at wpi.edu
Roadkill on the information superhighway.

On Mon, 29 Dec 1997, Paul Mozdziak wrote:

> I'm interested in separating things between 8-14 KB.
> 
> Any recommendations for optimal electrophoresis conditions to get
> optimal resolution?
> 
> I've been using SeaKem LE at 0.6% in TAE.
> 
> Thanks in advance.
> 
> 
> 
> 
> 




More information about the Methods mailing list