Plasmid DNA migration
hzhen at freeuk.com
Mon Apr 17 19:49:01 EST 2000
Nick Theodorakis wrote:
> the addition of EtBr will introduce (positive? - someone help me out
> here) supercoiled twists in DNA, and can dramatically effect its
> migration in agarose gels, which is greatly effected by conformation.
As someone else noted, the intercalation of EtBr first
relaxes the plasmid which was originally negatively
supercoiled. The plasmid can then runs slower due to the
relaxed conformation. When the incorporation of EtBr
increases to a critical point, the plasmid become completely
relaxed. Further intercalation of EtBr then introduce
positive supercoiling to the plasmid. At the maximum amount
of EtBr intercalation, the plasmid is again supercoiled, but
in the reverse direction (positive instead of negative).
There is a nice picture of the effect EtBr intercalation on
the conformation of plasmid and the sedimentation velocity
in the following article:
Bauer et al, Scientic American 1980 Jul;243(1):100-113
> also seem to recall an assay for topisomerase activity that used this
> effect to measure changes in supercoiling on a plasmid subsrate.
> Nick Theodorakis
> nicholas_theodorakis at urmc.rochester.edu
> Sent via Deja.com http://www.deja.com/
> Before you buy.
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