Reference: Zeng F and Quintana DG (2021) High-Copy Yeast Library Construction and High-Copy Rescue Genetic Screen in Saccharomyces cerevisiae. Methods Mol Biol 2196:77-83

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Abstract


High-copy rescue genetic screening is a powerful strategy for the identification of suppression genetic interactions in the model eukaryotic organism Saccharomyces cerevisiae (budding yeast). The strain carrying the mutant allele of interest is transformed with a genomic library cloned in a high-copy plasmid. Each clone carries a genomic fragment insertion of around 10 kb, typically containing one to three complete genes under their own promoters. The high-copy vector favors the accumulation of high levels of the corresponding protein, aimed at suppressing the mutant phenotype. Typically, high-copy genetic screens select for viable clones under conditions restrictive or lethal for the query mutant strain. Here, we describe in detail the procedure to generate a high-copy genomic library and a protocol for rescue genetic screening and identification of the suppressor clones.

Reference Type
Journal Article | Research Support, Non-U.S. Gov't
Authors
Zeng F, Quintana DG
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