Reddish Yeast in 2-Hybrid System? Help?

Heidi Moss hmmoss at MAIL.MED.CORNELL.EDU
Tue Jan 31 10:35:29 EST 1995


	Using the Brent 2-Hybrid System I have so far successfully cloned 
by bait into the DNAbd plasmid, transfected EGY48 w/ pSH18-34, checked 
for self reactivity, done a western to detect presence of the fusion 
protein, proceeded to transfect lib.plasmid containing a Hela cell lib, 
made glycerol stocks and proceeded to screen for protein interaction by 
plating onto Gal/Raf -His, -Ura, -Trp, -Leu plates.  For curiosity I also 
plated the same cells onto Gal/Raf -H,-U,-T and Glu -H,-U,-T.  The yeast 
colonies that grew the next day were reddish in color, where as the yeast 
on the Glu -H, -U,-T were white and present in high density as expected.  
Most interesting was the Gal/Raf -H,-U,-T plate contained a lawn of white 
colonies as well as a smaller number of large reddish colonies.  In all 
the plates the number of reddish colonies was the same so I assumed that 
these were in fact yeast that were growing on -Leu plates because of some 
relevant protein interaction.  Subsequently I have tested for Gal 
dependence and no color change was observed.  What is going on?  Can 
anyone help to explain these results? I would be very grateful.

						Kerry




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