high molecular plasmid DNA
(by aawara from pontiff-playground.org)
Fri Jan 11 22:52:25 EST 2008
In <mailman.153.1200074433.2451.methods from net.bio.net>,
Dr. Hiranya S. Roychowdhury <hroychow from nmsu.edu> wrote:
> Well, this has never been suficiently explained, nor do I know of any
> specific investigation into this. I tend to believe that the high MW
> species may exist as a complex of bacterial chromosome and other proteins.
> Usually, a clean plasmid prep does not show the HMW bands. It also has
> nothing to do with the conc. of the agarose gel.
Excessive treatment with alkali will cause this to appear. When nicked
DNA denatures, it creates a fast-migrating single-stranded circle that
is "undigestable" by restriction endonucleases. The other product is
linear ssDNA that tends to aggregate (by partial base-pairing) into
large complexes that migrates near the well of agarose gels.
If you treat minipreps with a single-strand specific nuclease like
S1 or mung-bean nuclease both aberrant species (hmw ssDNA, and fast-
migrating ssDNA) will disappear.
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