Magic minipreps--DNA don't cut

Julian Dow GBAA02 at udcf.gla.ac.uk
Sat Jan 15 06:52:46 EST 1994



> >In article <1994Jan13.171215.1 at samba.cnb.uam.es> mrege at samba.cnb.uam.es writes:
> > 
> >   Has anybody had any problem with Promega's Wizard Minipreps?
> >   In my hands the DNA looks well, but it is difficult to cut.
> >   Does  anybody know how to overcome this? Maybe washing the column twice?
> > 
> >   Thanks,
> >   Josefina
> >Yes, people in my group have had exactly this problem -- the DNA looks
> >great on a gel, but won't cut with certain enzymes (but will cut with
> >others).
> 

We've had problems with both mini and maxi preps. However, most of it could
be traced to not taking off ALL the supernatant when you first pellet the
cells. Given that the Promega Resuspension, lysis and neutralising
solutions are effectively IDENTICAL to the Maniatis 1-2-3 prep, we tend to
give things five minutes or so to work, and to use the neutralising
solution ice cold as per the 123 protocol. We also warm the TE to 70 C if
it's a big plasmid, and I'm tending to use distilled, rather than TE, to
elute. Generally, though, I'm a big fan of these preps. 
 I'm particularly impressed at how easy it is to elute a band from  a DNA
gel. Use ordinary, TBE / hmp agarose, nothing fancy. Cut the band out, pop
it into a 0.5 ml eppi, sling it in a PCR machine/ heating block till it
melts then pipette it into 1 ml of magic DNA clean up resin and buzz. Far
easier than electoeluting or glassmilk and very good yields!
-- 
 - Julian Dow



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