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Circular DNA & EthBr in electrophoresis

Mr. I.S. Viney iviney at hgmp.mrc.ac.uk
Tue Dec 12 07:35:32 EST 1995



On Tue, 5 Dec 1995, Bart Frank wrote:

> Can someone refer me to data on the migration of circular molecules
> in agarose gels with and without the gels being prestained with
> ethidium bromide?  We see a series of bands of slightly differing
> size when a 2 kb insert is ligated to itself in gels stained 
> after electrophoresis, but a predominately stained band
> if we place similarly treated samples in empty lanes of gels which
> had previously been stained before (& then after electrophoresis).
> Is this an expected anomaly?
> Thanks,
> Bart Frank
> 

I think this is an expected anomaly, the forms which migrate slightly
differently are topoisomers.  Pretreatment with EtBr destroys the
differences in mobility, maybe someone else (better versed in DNa
topology!) can explain this clearly.  The same thing is seen with
preparations of M13 RF DNA and can lead to confusing band patterns in
uncut DNA samples if the gel doesn't contain EtBr before electrophoresis,
I guess the products of a ligation reaction are similarly afflicted. 

Hope this sounds plausible!

Ian Viney
Imperial College
  



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