anichol at larry.cc.emory.edu (Ainsley Clare Bayliss) wrote:
>> Does anyone
> have experience with plating yeast on Xgal? Can I increase the acidity
> of my media somewhat without losing the activity of the Xgal? Is there
> any way to speed up the color change? Also, does anyone know the
> specific activity of B-galactosidase? What specific activity is
> necessary for yeast to turn blue?
Sometimes I think my entire thesis was beta gal platings....
If you are plating on XG directly, rather than using filter lifts, then
you will need to adjust the pH of the media for maximal sensitivity.
Check Lenny Guarente's MEth. Enzymol. paper from years back on using
beta gal fusions in cerevisiae for a recipe (at least, I THINK that's
the reference).
If you measure beta gal activity in Miller units (that is, units/OD
of liquid cell culture) and compare that to plate assays,
then XG plates and a sharp eye can pick up anything
from about 2 units on (GAL, for comparison, induces to the 1000-range
as I recall). Beta gal is stable so you tend to accumulate it and
colonies will slowly become blue with time, unless they are not
expressing beta gal at all.
hope this helps
susan
susan_forsburg at qm.salk.edu
