Referring to sequencing with the ABI 373A sequencer we have big problems.
Twice this year (March and October) our sequencing results have been
horrible for a month at a time, and it appears to be starting again.
That means that we get very low signal heights and the readable sequence
is about 0 - 50 bp. Both times, the problems have
occured suddenly after months of very good results (we changed none of
the reaction conditions). We use the dye terminator cycle sequencing kit
with the recommended programm from Applied Biosystems. To try to get at
the sequencing problems we have replaced ALL our solutions, we tested a
fresh premix solution, we used DNA preps which had shown good sequencing
results a week before, we have used primers freshly diluted from our
stock solution (4 different ones, bought at different times), we have
used phenol and CTAB precipitation but nothing helped and our sequencing
results were as bad as before. We checked the machine and the gel with
a sequencing reaction made by ABI and this works fine. Both previous times,
the problem solved itself and we don't know why. This is costing much
time, money and frustration and we would love a solution. So now we don´t
know what to do. Could somebody please give us some suggestions.
With my thanks in advance,
P.S. We wrote ABI (technical service) twice regarding the above and
they have not responded.