Rhodamine dUTP and capillary eletrophoresis

Edward Weiss via methods%40net.bio.net (by edward.weiss from utoronto.ca)
Tue Dec 11 15:51:35 EST 2007


Hi all,

Just wondering if anyone's run into this problem or has any suggestions 
for how to remedy it. We've started labeling select PCR products with 
rhodamine-dUTP and running them on our ABI 3730xl. So far, so good, at 
least when the fragments are run right after the PCR. However, we've 
found that if we try using a restriction enzyme or a 
mismatch-recognizing enzyme (CEL I) on these labeled samples, the 
intensity on the ABI is so weak as to be almost negligible. These same 
samples look fine when run on an agarose gel, so I'm guessing that for 
some reason the labeled products are not making it into the capillaries, 
perhaps to the rhodamine binding something or another. Any ideas about 
how we can circumvent this?

Thanks,

Ed

-- 
Edward S. Weiss
Research Technologist
Genetic Analysis Facility
The Centre for Applied Genomics @
The Hospital for Sick Children
Room 14-601S, East Tower
101 College Street
Toronto, ON M5G 1L7
Canada

416-813-8642
edward.weiss from utoronto.ca



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