l.adams at garvan.unsw.edu.au
Wed Nov 27 15:51:04 EST 1996
I am also having problems with D5S392.
We are using an ABI377 sequencer, and the primer is labelled with FAM. I
also get some non-polymorphic bands in this region. On some occasions I do
get a product in the correct size range (83-117 bp), although the
fluorescent signal is very low (around 60-100). Furthermore, I get what
appears to be a second microsatellite at approximately 190-200 bp. This
second set of bands has a much higher level of fluorescence - I had
initially mistaken these for the "real" bands before realising that it did
not fall anywhere near the expected size range.
The conditions we use are a 55 degree annealing temperature. We use TaqGold
and a 2.5 mM concentration of MgCl2 (which we use for all primers, but
which I am considering lowering to see if this helps). Our primers are the
same as those of Flemming, although we have an extra base added on the
forward primer, and two bases on the reverse primer (on the 3' end in both
I would also like to hear if anyone is having success in using this primer.
Linda Adams, Ph.D.
Garvan Institute of Medical Research
St Vincent's Hospital Tel: (02) 9295 8288
Sydney NSW 2010 Fax: (02) 9295 8281
Australia email: l.adams at garvan.unsw.edu.au
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