Yeast Plasmid DNA isolation

Frederik Boernke boernke at nospam.ipk-gatersleben.de
Fri Jun 19 06:26:07 EST 1998


gmclab221 wrote:
> 
> Hello,
> I was wondering if anyone has done restriction enzyme analysis on
> plasmid DNA isolated directly from yeast by the rapid mini-prep method
> (i.e. breaking buffer, beads and phenol/chloroform/IAA).  I am trying to
> clone particular inserts into a plasmid of interest by homologous
> recombination in yeast; however, my background is considerably high so I
> was hoping to do a rapid initial screen and then go into E.coli after I
> found the correct plamid containing yeast colony. Does anyone know what
> the yield of DNA is generally for these preps and whether it is possible
> to detect a cloned insert by restriction analysis with these preps?
> Thanks you so much for you help.
> Any advice woudl be greatly appreciated.
> Nancy
> nayers at cctr.umkc.edu

Hi Nancy,

I doubt that restriction analysis would get you any further in this
case,
but can't you use PCR to test for the presence of your insert by the use
of insert flanking primers? You could easily  avoid making loads of 
plasmidpreps by just PCR directly from the colonie.

Good luck
Ricky

******************************************************************

Frederik Boernke
Research Group of  Molecular Plant Physiology
Institute for Plant Genetics and Crop Plant Research (IPK)
Corrensstr. 3
06466 Gatersleben
Tel.  039482 -5 321
Fax. 039482 -5 515
http://www.ipk-gatersleben.de




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