Dissolving plasmid DNA after precipitation

Lou Cantolupo CANTOLUPO at GENVEC.COM
Fri Jan 24 08:57:53 EST 1997


How big are your plasmids?  The bigger the DNA, the harder it is to get
into solution.  A plasmid 2-10Kb should have no problem going back into
solution quickly, but DNA (such as adeno, 35Kb) can take up to a day to
do so.  

I (and quite possibly 99.9% of all molec.biol's out there) follow the
typical protocol of a 1/10 NaOAc precip.  However, my first precip is
with 1 volume isopropanol, then the 70% EtOH wash--just one.  This
shouldn't make a difference.  If your DNA gets too dry, yes, it may not
resuspend, but I highly doubt this is salt related.  You can also try 5M
PotassiumOAc and 10M NH4OAc.  Check Maniatis/CP or whatever for volumes
and pH's on the salts; I don't remember.  Any residual EtOH in your
pellet will affect it's ability of going back into solution as well.  I
tend to dry my smaller DNA's in a speed vac to get rid of it all.  

Lou Cantolupo
Dept. Molecular Virology
GenVec, Inc.
CANTOLUPO at GENVEC.COM




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