Yeast maxi plasmid preps?
nc1 at acpub.duke.edu
Fri Mar 7 11:50:37 EST 1997
In article <331FFF2D.6E48 at esbs.u-strasbg.fr>, Kathy Baynton
<baynton at esbs.u-strasbg.fr> wrote:
> I was wondering if anyone knows if it is possible to produce maxi
> preps of plasmids from yeast that are relatively clean and in
> respectable quantities (ie. using 2mu-based origin plasmids) as is
> done in E. coli? (ie CsCl purification, Wizard). I am already aware
> of plasmid isolation preps from yeast (Hoffman and Winston I believe
> is the reference), but these preps are pretty dirty, the plasmid is
> not really "purified" and we always end up having to re-transform into
> E. coli in order to recover plasmids that are "clean enough" to do
> sequencing, cloning etc. I am also aware of PCR-based sequening that
> can be done on yeast-isolated plasmids but for what we're thinking of
> doing, I'm not sure that this would be an appropriate method.
As a person who has always been wondering if your kind of idea would be
possible, I don't know of any reference or researchers who could do
plasmid maxi prep, or even mini prep, directly from yeast.
The trick that make us isolate plasmids from bacterial chromosome is that
the latter is attached to membrane while the former is not; this is no
longer true for yeast! As you mentioned, Hoffman and Winston method is to
release plasmid, but not to isolate it from yeast cells.
Please let me know if you find any good idea.
Namjin Chung, who knows little and asks a lot!
Program in Molecular Cancer Biology
Duke University Medical Center
Durham, North Carolina
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