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Titering Gal1 promoter

Jon Schmuke jjschm at ccmail.monsanto.com
Thu Aug 31 17:02:03 EST 1995


In article <424qdb$opj at swsu65.swmed.edu>, Mark Walberg
<walberg at simmons.swmed.edu> wrote:

> Regarding titering the GAL1 promoter with mixtures of glucose and 
> galactose:  I think that this is going to be very difficult to do in a 
> controlled way.  In such mixtures I would expect a diauxic growth 
> pattern with consumption of glucose first, then as glucose concentration 
> falls the GAL1 promoter will become activated.

I suspected this would be a problem.  In the first experiment I used 4%
raffinose in the presence of increasing amounts of galactose.  Invitrogen
suggested using raffinose as a non-repressor/inducer source of carbon. 
Since the gene is essential, the increasing level of galactose won't
enhance growth unless the enzyme level is increased via the GAL1 promoter. 
The cells wil not grow in raffinose alone but grow at the same rate with
0.2% or 2% galactose.  If the enzyme titer need only be a few copies or if
the GAL1 is an "all or nothing" promoter this might make sense.  I was
hoping that perhaps I could titer it better by repressing with glucose, but
perhaps switching to another promoter would be better.  CUP1 was one
suggestion.

Thanks for all your help,

Jon

***************************************************************
Jon J. Schmuke                       jjschm at ccmail.monsanto.com
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