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What do you suggest for yTHC troubleshooting?
Two references may help: one detailing the creation of the collection and the other one detailing its use. For this collection, each strain was qualified by its growth phenotype on YPD with 10µg/ml doxycycline.
Exploration of Essential Gene Functions
via Titratable Promoter Alleles, Mnaimneh et. al., Cell, Vol. 118, 31–44, July 9, 2004, Copyright ?2004 by Cell Press
Functional Discovery via a Compendium
of Expression Profiles, Hughes et. al., Cell, Vol. 102, 109–126, July 7, 2000, Copyright ?2000 by Cell Press
Without doxycycline in the media the Tet-promoter is fully activated. Addition of doxycycline in a titratable manner allows for down regulation of the promoter until the gene of interest is no longer expressed at detectable levels. The optimal condition for each strain needs to be determined empirically. It is very important to perform serial dilutions of both the yeast and the doxycycline. If you have too heavy an inoculum of yeast, the expression of the gene or protein may be too great to allow for any reduction to be detectable.