ABI 3100/3700 genetic analyzer: substitute NED dye with TAMRA?

Deanne Bell dbell at qnis.net
Fri Dec 1 17:11:26 EST 2000


Hi
I don't know if the filters are the same on your machines but I am using
the ABI 310.  And on my machine, TAMRA is detected as red if you are using
the Virtual filter set 'C'.  TAMRA is detected as yellow using virtual
filter set 'A'.  When we started using our ABI 310 for AFLPs, Applied
Biosystems put out a kit of primers using TAMRA as the yellow dye.  They
then switched over to using NED as the yellow dye in the primer kit.  I
think I remember them saying that the TAMRA had more bleeding over into
other colors than the NED.  Either which way, I never got the yellow
labeled primers to come off with any height on my electropherograms - be it
NED or TAMRA.  I am working with plants and it may be that these primers
just don't amplify much in plants (although ApBiosystems claims to  get
amplification in plants with these primers).  I have a feeling it is more
likely that the dye just doesn't emit as well as the other dyes such as
JOE, FAM & ROX.  I would avoid using a system relying on NED or TAMRA as my
markers / standards.  See if you can and IMHO, switch over to a set-up
where you use ROX for your markers / standards.

Deanne Bell
Molecular Markers Lab Technician
USDA Agricultural Research Service
2021 S. Peach Ave.
Fresno, CA 93727
e-mail: dbell at qnis.net
phone: 559 453-3170
fax: 559 453-3088

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From: "Marcia Macdonald" <mmacdonald at xenongenetics.com>
To: methods at hgmp.mrc.ac.uk
Subject: ABI 3100/3700 genetic analyzer: substitute NED dye with TAMRA?
Date: Friday, December 01, 2000 11:05 AM

Has anyone else tried to avoid buying NED-labeled oligos from ABI by
substituting TAMRA-labeled oligos?  We just did a quick experiment on our
3100 and found that the TAMRA was detected as yellow peaks just like NED! 
Like the other dyes, there was a small amount of leakage into the other
colours when the PCR product was over-loaded, but nothing that will
interfere significantly with pooling multiple markers  We still have to
compare the same marker amplified with both TAMRA and NED dyes to see how
their mobilities compare (i.e. if the alleles are labeled as the same
size), but that shouldn't prevent us from ordering new markers in the TAMRA
dye rather than the NED.  Just thought this might interest anyone
genotyping or synthesizing labeled oligos.

Marcia

Marcia MacDonald
Xenon Genetics, Inc.
100-2386 East Mall
Vancouver, BC  
Canada  V6T 1Z3
604-221-8423 fax


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