High-Copy Yeast Library Construction and High-Copy Rescue Genetic Screen in Saccharomyces cerevisiae

Fanli Zeng*, David G. Quintana

*Corresponding author for this work

Research output: Chapter in BookChapterResearchpeer-review

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.

Original languageUndefined/Unknown
Title of host publicationMethods in Molecular Biology
Pages77-83
Number of pages7
DOIs
Publication statusPublished - 2021

Publication series

NameMethods in Molecular Biology
Volume2196
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029

Keywords

  • 2 μm
  • Budding yeast
  • Dosage suppression
  • Genetic screen
  • High-copy genomic library
  • High-copy suppressor
  • Saccharomyces cerevisiae

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