dye labelled primer extension

Justin Powell jacp1 at mole.bio.cam.ac.uk
Mon Aug 28 13:54:38 EST 2000


Has anyone tried to do 5' transcriptional start mapping using fluorescent
sequencing technology rather than radioactive?  I am thinking along the
lines of using dye labelled primers and doing an RT primer extension using
one of the sequecing primers which would then be run in an adjacent lane
to the sequencing reaction on an ABI377?

I am wondering whether a) the signal from the primer extension would be
strong enough, b) whether the adjacent lanes run so that positions
correspond accurately and c) whether the analysis software can display the
two traces side by side or overlayed such that their positions correspond
(I am used to just getting the traces for individual lanes sent back from
the sequencing facility we have, and wouldn't know how to line one up with
the other, there seem to be coordinates of some sort displayed above the
trace are these gel positions?).

Dr.  Justin Powell

Dept. of Genetics
University of Cambridge
Downing Street
Cambridge
UK
CB2 3EH
Tel.(01223) 333985
Fax.(01223) 333992






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