SDS-PAGE problems

Edward Theodore Michelini tedm at darkwing.uoregon.edu
Mon Aug 14 17:52:08 EST 1995


Fellow Bionetters,

   Recently I have been trying to do SDS-PAGE on a thermophilic (80¡C) 
bacteria which is known to have some very odd, and copious, membrane 
lipids. My standard SDS loading buffer with BME/SDS seems to lyse them a 
bit when boiled for ~10min but the resultant gels have very little info 
under 40kD, perhaps a bit blurry. Is this due to incomplete denaturation 
of the proteins, with smaller, tougher ones less likely to take on the 
SDS? Or has it been seen with large amounts of lipids? I have been told 
that urea in the stacker will possibly aid in the denaturation of the 
proteins, and maybe I will sonicate the cells to assure lysis, but can 
the lipids be removed with a CHCl3 exraction?  Any insights would be welcome.

Thanks in advance,

Ted Michelini
Institute of Molecular Biology
University of Oregon
tedm at darkwing.uoregon.edu



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