help on DNA sequence problem,anxious

Carnac carnac at realitys-edge.com
Tue Dec 31 00:07:32 EST 2002


<shelley025 at yahoo.com> wrote in message
news:20021230175946.31737.qmail at ww02.hostica.com...
>
> Help on DNA sequencing:urgently
>
> Use T7 primer, big dye terminator chemistry. However the signal stops at
~300BP. Maybe there is secondary structure lies in that region.  I tried
several ways to solve this problem. Adding DMSO, increasing denature
temperature, using different primers. But it still does not work.
> Who met this same problem before? Can you give me some suggestion?

Is your DNA clean?  No protein remaining?  Have you tried to run the DNA out
on a gel first to make sure your DNA length is good?  The fragment isn't
methylated?  Tried increasing the time for the elongation phase?






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