Do you need supercoiled pDNA for ds-sequencing?
jow at helix.nih.gov
Tue Nov 1 17:11:40 EST 1994
In article <MURIANAP.30.000C1D5C at FOODSCI.PURDUE.EDU> Pete Muriana,
MURIANAP at FOODSCI.PURDUE.EDU writes:
>Now the question - the DNA sequencing facility representative indicated
>we need to have **supercoiled** plasmid DNA to sequence from. Is this
>and why wouldn't a linear double-stranded fragment work? Am I missing
I can only speculate. My experience agrees with your sequencing facility
representative. I have always had better luck sequencing intact, mostly
supercoiled plasmid than with linear double-stranded DNA.
Maybe there is less likelyhood of reannealing of the strands when coiled
DNAs are snap denatured?
More information about the Methods