PCR sequencing

Victor JONGENEEL vjongene at ulci20.unil.ch
Tue Feb 9 02:42:34 EST 1993



There have been several postings recently requesting methods for direct
sequencing of double-stranded linear DNA molecules.  This is perfectly possible
as long as two things are kept in mind: (i) the annealing time for the sequencing
primer should be *very* short to keep the DNA strands from re-aneealing; (ii) the
elongation and termination reaction times should be shorter than those used in
sequencing supercoiled or single-stranded DNAs, for the same reasons.

A short protocol can be found in Casanova et al., Nucl. Acids Res. 18:4028, 1990,
and a much expanded version with detailed protocols in a book to be released soon
by Humana Press: "Methods in Molecular Biology: DNA sequencing", Ed: H. Griffin.

Good luck to all!

Victor Jongeneel
Ludwig Institute for Cancer Research



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