dsDNA cycle sequencing - HELP

Paulo Magalhaes pamaga at biobase.aau.dk
Wed Jul 15 08:31:14 EST 1992


I'm trying to set up a "dsDNA cycle sequencing" protocol,
using a clean dsDNA PCR product as template, an unlabeled
oligo, a mixture of the 4 dNTPs, 35-SdATP and ddNTPs.
My A and C lanes show very faint bands, especially for
the longer fragments, which I presume is due to a bad
d:ddNTP ratio.
Can someone help me save many hours of trials?
I don't want to use 32-PdATP.
A detailed protocol is most welcome.
Thank you.

Paulo

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Paulo Magalhaes
Rigshospitalet
Copenhagen, Denmark

email: pamaga at biobase.aau.dk

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