Full genome repressible yeast library that can be tailor-made for specific screens

Despite decades of research we still do not know the function for the vast majority of eukaryotic genes. The yeast Saccharomyces cerevisiae has been a strong model cell for functional genomics underlying the discovery of many conserved gene-functions. One prominent tool in functional discovery is the yeast knock-out library, which enables to screen the yeast genome for phenotypic effect on a desired biological process. However, the library, created over 20 years ago, suffers from major shortcomings such as that it naturally lacks essential genes, has been depleted over the years of slow-growing strains, has accumulated many suppressor mutations and lack of representation of any newly identified genes. To overcome these hurdles and create a new and improved loss-of-function library, we utilized the SWAp-Tag (SWAT) method, developed in our lab, to create a new full genome library with repressible alleles by incorporating the newly developed Z3 inducible promoter in which all the proteins are also N-terminally tagged with mScarlet. This library can used to screen for specific pathways or phenotypes by re-making it with any given genomic modification of choice, a process which takes three weeks. I will show how we utilize these properties for discovering new organelle and protein trafficking related genes, using genomically encoded fluorescent organelle markers.