Need help - black dots on DNA sequencing gels?

Robert E Minto reminto at jhunix.hcf.jhu.edu
Tue Sep 6 09:34:30 EST 1994


I am in need of some help removing interfering black dots from my 
polyacrylamide sequencing gels.  The gels are prepared by the U.S. 
Biochemical Sequencing support guide protocol which contains 8M urea (and 
appears to be identical to Maniatis, 6% acrylamide, 19:1 
acrylamide:bisacrylamide).  The gels contain black dots which are 
concentrated at the top of the sequencing gel and are ONLY in the lanes.  
They actually interupt the lanes, suggesting particles in  the gel.  The 
gels even contain dots when poured from fresh reagents (Mol. Biol. 
grade), with aspirator-degassed solutions carefully swirled with TMED and 
APS to avoid bubbles, and polymerized for 2 hours before using 
immediately. Both vertical and horizontal (capillary action) pouring 
techniques give the same result.  The buffer is 1 x TBE.  The operating 
temperature is ca 40-45 degrees at 60W power.

Any suggestions as to the cause of this irritating phenomenom would be 
appreciated.   

Thank you.  Robert Minto  (reminto at jhunix.hcf.jhu.edu)


-- 


Robert Minto
reminto at jhunix.hcf.jhu.edu



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