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.
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Acknowledgments
This work was supported by the National Natural Science Foundation of China to FZ (grant No. 31801039) and by the Ministry of Economy and Competitiveness of Spain (MINECO) and the European Regional Development Fund (FEDER) to DGQ (grant BFU2015-68493-P).
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Zeng, F., Quintana, D.G. (2021). High-Copy Yeast Library Construction and High-Copy Rescue Genetic Screen in Saccharomyces cerevisiae. In: Xiao, W. (eds) Yeast Protocols. Methods in Molecular Biology, vol 2196. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0868-5_7
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DOI: https://doi.org/10.1007/978-1-0716-0868-5_7
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Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-0867-8
Online ISBN: 978-1-0716-0868-5
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