Yeast two hybrid contamination

Matt Benson mabenson at
Tue Mar 10 05:15:48 EST 1998


I've now done about 4 yeast 2-hybrid screens and every time after 5 days
incubation at 30C am getting small, brownish, glassy colonies which
absolutely stink.  (And no positives!) These look nothing like my yeast,
I've checked out my bait strain extensively for self activation and using a
known interactor (which worked perfectly ), each screen has used fresh media
and colony PCR on the small, smelly colonies show that they do not contain a
prey plasmid.  I'm using a mouse brain cDNA library from Origene with the
Hybrid Hunter system from Invitrogen.  Two of the screens have been
performed with the Gietz method and two with the Clontech manual variation. 

I am now getting very frustrated!

Any ideas greatly appreciated



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