fluorescent detection: microsatellites (help)

ParentE at dfo-mpo.gc.ca ParentE at dfo-mpo.gc.ca
Tue Sep 4 09:33:55 EST 2001


Hello all.

We have a problem with fluorescent primers and would need advice. We have
been doing microsatellites work on snow crab for awhile now and never had
any (not much) problem using radioactively labelled primers.  We recently
purchase an ABI 310 automatic sequencer and are in the process of quitting
radioactive work for fluorescent primers.  The problem starts here. For some
loci we didn't had any problem, but in some case we can't get any signal.
The weird thing is that we tried to labelled one primer with radioisotopes
and use the fluorescent primer as reverse. We get a good signal on
acrylamide. When using the same primers set but without radio labelling we
don't get any signal on the sequencer. Should we try to re optimise the PCR
or is there a simple explanation to our problem?

I must admit that I'm baffled by all that and need advice very quickly


Thank you in advance for your help


Eric Parent

Maurice Lamontagne Institute
Fisheries and Oceans Canada
850 route de la mer
C.P. 1000
Mont-Joli, Qc
Canada
G5H 3Z4

parente at dfo-mpo.gc.ca
http://www.qc.dfo-mpo.gc.ca/iml/

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