Sequencing problem, HELP!!

[Glen Shearer]

Zainul Zain (zainul at KB.USM.MY) wrote:
: Greeting to all bio netters,

: I recently had a problem with my sequencing gel. When I load my sample 
: they did not run in a straight line as they should be, instead, they 
: dispersed from the straight line.

:                      
:                      _     
:                  _       _
:                _            _   This is how they looks like on running gel!

: This happen to me when I change the acralymide soln. to the new long 
: ranger solution. I prepare 6% gel and followed exactly the protocol 
: provided by the manufacturer.

: Please advice!!!

   We recently had the same problem with 'frowns'
The problem turned out to be GSE (Grad Student Error).
He had decided to put a clamp on the bottom, center of the glass
plates (without a bottom spacer) before pouring the gel.
Thus the gel became thinner than 0.4mm toward the bottom center.
Thus the frown.....
   The fix:  don't clamp the bottom of the plates.  In fact, we
now use only 0.4 mm side spacers, a strip of chromatography paper
instead of a bottom spacer and do not tape the plates at all.  Just
put three spring clips on each side.  Quick, no leaks.