Fluorescent sequencing of high(er) MW dsDNA

krishnan at BORCIM.WUSTL.EDU krishnan at BORCIM.WUSTL.EDU
Tue Mar 29 08:59:01 EST 1994


Hi folks!
To my reading, most of the fluorescent sequencing seems to be with single-
stranded DNA template and comparatively less with small plasmid templates, 
or PCR fragments.  What is the netland experience on sequencing double-
stranded templates of  varying  sizes (say range of 5-25 kb)?  Dugaiczyk 
et al (1992) NAR 20:6421-6422 reported that linearizing cosmid sized templates 
improved accuracy.  Has this been the general experience too for ds DNA 
or is this required only when dealing with complex (=cosmid sized large) 
templates?  Also, could somebody tell me whether using dye-primer or 
dye-terminator chemistries makes a difference in the length and/or accuracy of
 read obtained, particularly when directly  sequencing ds DNA?
I shall be glad to post a summary.
Much appreciated.
Raja



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