PCR on whole yeast cells (fwd)

Robert Preston rapr at MED.PITT.EDU
Tue Jun 14 08:44:14 EST 1994


> In article <2t7ks6$a9a at mserv1.dl.ac.uk> Dr. I. Dunham, idunham at crc.ac.uk
> writes:
> >
> >Incidently Eric Green has recently found that this colony PCR does not
> work 
> >reproducibly in thermocylcers with a heated top plate if you omit the 
> >mineral oil.
> 
> Does anyone have a theory as to why that would be?
> 
> Dave Knorr
> 
Sure. (talk is cheap!) There has been talk on the net about how it seems
that, in some cases, mineral oil is a sink for inhibitory traces of
detergents in some DNA prep. protocols.  Yeast is eukaryotic, meaning it
has (relative to the average prok. vector) gobs of intracellular membrane-
bound organelles (among other things) and membranes have phopholipids (i.e.,
bio-type detergents) and may inhibit PCR, unless soaked up by mineral oil.
The prediction is that chloroform extraction of the colony would make
mineral oil unnecessary (although still more convenient, maybe, than
the proposed extraction)

Rob Preston
rapr at med.pitt.edu




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