dye labelled primer extension

Arnoud van Vliet avvliet at knoware.nl
Mon Aug 28 15:25:10 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?

Dear Justin

we have been thinking about this as well. I have seen a reference where they
used fluorescent primers and an ALF sequencer for primer extension. The
references are:

Grass, G., C. Große, and D. Nies. 2000. Regulation of the cnr cobalt and
nickel resistance determinant of Ralstonia sp. strain CH34. J. Bacteriol.

Große, C., G. Grass, A. Anton, S. Franke, A. Navarrete Santos, B. Lawley, N.
L. Brown, and D. H. Nies. 1999. Transcriptional organization of the czc
heavy metal homoeostasis determinant from Alcaligenes eutrophus. J.
Bacteriol. 181:2385-2393

If you get the primer extension running, we'd be very interested in the
protocol, and maybe you can post it on the newsgroup?

Hope this helps

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