Glucose reppression of gal promoters

Desrochers Normand desrocn at ERE.UMontreal.CA
Mon Dec 19 15:35:46 EST 1994


coutave at rockvax.rockefeller.edu (Elias E. Coutavas) wrote:

>A friend has an essential gene behind a gal promoter.  Shift to glucose
>does not kill the cells.  Is this likely suppressor mutations or
>readthrough?  No RNA can be detected on glucose.
>Respond here or by email to him directly at aitchij at rockvax.rockefeller.edu
>Thank you.


	We work with an essential gene too and we used to make plasmid 
shuffle to be sure the plasmid is expunged. We just had to put the gene 
on an URA3 plasmid and transform in a diploid strain that got a knock out 
of the gene. We dissected and kept the haploid cells that got the 
knock-out (at this step, the plasmid must be necessary for the cell 
viability). Finally you just have to streak this strain on a 5-FOA media. 
Generally, no growth should be detected. If the reason of your friend's 
problems is a high stability of the protein, maybe it will not work.
Tell him good luck!


      ********************* DESROCHERS, Normand **********************
      **************** desrocn at alize.ere.umontreal.ca ****************
      ************** dep. Microbiologie et Immunologie ***************
      ******* Universite de Montreal (514) 343-6111 poste:1432 *******



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