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377 base spacing problems

Frazer Murray Frazer.Murray at bbsrc.ac.uk
Tue Jun 8 15:54:26 EST 1999


Has anyone had problems with the base-spacing when analysing their 96 lane
sequencing gels on the PE 377? I am running 48 lane gels fine with spacings
about 10 or 11 but when running a 96 lane with the same conditions the
spacings are given as defaults of red 9 and the sequences are unusable. It
is as if the gel is running too slow. Do I need to increase the voltage
that the gel is run at when running 96 lanes? At the moment it is at 1650V.
I am using millipore collumns with G75 sephadex to clean up my termination
reactions (big dye). Some people have suggested there is salt passing
through into my sample but the gel image lookes fine and straight except
for the very last few bases where it pinches in. I believe that salt would
result in a more dramatic pinching.  
Any ideas would be most welcome

-- 
Fraze Murray
Roslin Institute




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