timed induction of gal1 promoter

Soon A. Lee s.lee at auckland.ac.nz
Thu Feb 29 15:44:37 EST 1996


Dear net readers,
I wonder if you could help me.  My problem is this - I want to do northerns/growth curves on strains overexpressing certain genes from the gal1 promoter.  The overexpression decreases the growth rate, meaning I can't harvest the strains all at once at log phase for analysis (if I grow from scatch on galactose).  I would like to induce the promoter at say OD600 0.3-0.4, then allow four hours more growth for induction before harvest.  What is the best way to do this?  I have heard of growing on glycerol/lactate medium followed by galactose addition at the appropriate time (anyone have a description of glycerol/lactate synthetic medium?).  However, for certain reasons I would prefer to use a fermentable substrate.  Could I make medium with a low glucose conc, with normal galactose, and rely on depletion of the glucose for induction?  How much glucose should I use to see depletion by about early log phase?  Any tips would be much appreciated.
Cheers,
Colin MacDiarmid
University of Auckland 
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