Acrylamide electrophoresis of DNA problem

Albert FAlbert at bilbo.bio.purdue.edu
Tue Oct 18 09:44:37 EST 1994


To the electrophoresis gurus of the world:

I have been utilizing acrylamide electrophoresis as a means of studying DNA
curvature and I've recently encountered a problem.  The addition of certain
drugs to my sample buffer will restore near-normal electrophoretic mobility
to the bent DNA fragments while not affecting the mobility of the non-bent
controls.  Here is the problem:  I have one highly bent DNA fragment that
smears upon the addition of any the drugs regardless of the DNA purity or 
electrophoretic conditions.  On gels where greater than 100 DNA fragments
(both bent and non-bent) are highly resolved at all drug concentrations,
this one particular booger continues to smear downward.  Without any drug
the DNA band looks just fine.  It is suggestive of a heterogeneous
population or some DNA/drug affinity problem.  Gel systems which include
the drug in the gel matrix and resevoir buffers have the same results and
the literature indicates that these drugs have extremely low dissociation
constants.  Recloning the fragment has not helped.  Mutants of the DNA
sequence (point mutations that affect the DNA structure) display the same
selective 'smearing' response to the drugs.

Question:  Can anyone tell me more about the electrophoretic properties of
DNA?
           Why is this effect selective?  

	
Any comments or suggestions will be greatly appreciated,
											                             Fred Albert
																																								FAlbert at bilbo.bio.purdue.edu



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