In article <5ct2rc$f64 at net.bio.net> Muhammad A. Lodhi, lodhi at cshl.org
writes:
>Dear Friends,
>>Would someone like to share with me his/her experience of sequencing
>through a region of ~22 As in Arabidopsis. I have tried different
>chemistries from ABI and Amersham but both fail to produce any good
>sequence. It produces very nice and clean sequence till the end of the A
>stretch but fails right after that. I have not tried using anything
>different in the reaction yet. Recently I heard about Dynamic ET poly T
>primer kit of Amersham but haven't tried it either. Any advice of any
>sort
>will be appreciated.
>Cheers!
I have heard that an oligo dT primer is best for this (avoiding stutter
which
gives garbage after the As) but I got through the A stretch I had with a
primer
that just bound very close to it. Alternatively, can you come in from the
other side? ALternatively again, Sequenase with dye terminators is much
more
processive than Taq, and so you might solve the stutter problem that way.
Hope this helps,
Anthony.
