Do you need supercoiled pDNA for ds-sequencing?

Pete Muriana MURIANAP at FOODSCI.PURDUE.EDU
Tue Nov 1 12:06:48 EST 1994


I recently spoke to a person with a DNA sequencing facility about isolating a 
plasmid restriction fragment to sequence from without the need to clone the 
fragment (although the cloning of a plasmid fragment is a simple task).  Since 
we have mega-quantities of the plasmid, we thought of gel-eluting the desired 
fragment; we have a gene-based probe encoded on the fragment (already 
demonstrated by Southern blot).

Now the question - the DNA sequencing facility representative indicated that 
we need to have **supercoiled** plasmid DNA to sequence from.  Is this true? 
and why wouldn't a linear double-stranded fragment work? Am I missing 
something here?

Regards, Peter
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*  Peter M. Muriana, Ph.D.             317-494-8284   TEL            *
*  Dept. of Food Science               317-494-7953   FAX            *
*  Purdue University                   murianap at foodsci.purdue.edu   *
*  Smith Hall                                                        *
*  W. Lafayette, IN  47907-1160                                      *
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