digesting agarose embedded DNA

Rob Jordan rjordan at u.washington.edu
Wed May 14 17:12:00 EST 1997


I will be digesting agarose embedded DNA soon and the protocol I have that 
comes with the PFGE system says it may be desirable to wash the plugs with 1mM 
PMSF after proteinase treatment, but before restricting.  I have a couple of 
questions:

1.  Is it really necessary?  I'll most likely be doing the restriction digest 
overnight and the proteinase treatment protocol calls for 4, 50ml washes.

2.  If it is a good idea, what solution should I dissolve the PMSF in?  The 
wash solution is just 20 mM Tris, pH 8.0, 50 mM EDTA.  Is it ok to just make a 
50 or 100mM solution in this or do I need to use a different solvent?  How 
long will the PMSF last in solution?  How should I store it?

Thanks in advance-
Rob
rjordan at u.washington.edu


Rob Jordan
Radiation Biology Laboratory
University of Washington Medical Center
Box 356069
Seattle, WA 98105



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