Question about plating yeast transformants directly on selective gal plates
Dr. Joe S. Mymryk
jm8v at AVERY.MED.VIRGINIA.EDU
Wed Feb 26 22:02:04 EST 1997
Recently, I was trying to transform a W303 derived yeast line with
plasmid and plate directly on selective media containing galactose. This
was necessary for technical reasons. This procedure yielded few if any
transformants when plated on SC-galactose, although good efficiency was
obtained if the other half the transformation mix was plated on
SC-glucose. Prior to Lithium Acetate transformation, I was growing the
cells in YP-glucose.
The lack of transformants on SC-galactose may be related to plasmid
establishment as transformants isolated from the SC-glucose plates will
grow when streaked on the SC-galactose plates.
So far I have tried two things that get around this problem.
1). If I growth the yeast for 16 hours in SC-glucose media in liquid
culture immediately after the plasmid transformation, yeast will grow
when plated on SC-galactose. This may give the plasmid a chance to
become established before the switch to galactose?
2) If I grow the yeast in YP-galactose in preparation for the
transformation, I can plate directly on SC-galactose plates.
My questions are:
1) Is it standard practice to grow yeast in YP-galactose prior to
plasmid transformation that will be plated on SC-galactose media?
2) Why is the transformation efficiency so poor on SC-galacotse if the
yeast are grown on glucose prior to plasmid transformation? Any idea of
Thanks in advance for your input,
Joe Mymryk, Ph.D.
jm8v at avery.med.virginia.edu
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