Systematic exploration of essential yeast gene function with temperature-sensitive mutants.

• Conditional temperature-sensitive (ts) mutations are valuable reagents for studying essential genes in the yeast Saccharomyces cerevisiae. We constructed 787 ts strains, covering 497 (∼45%) of the 1,101 essential yeast genes, with ∼30% of the genes represented by multiple alleles. All of the alleles are integrated into their native genomic locus in the S288C common reference strain and are linked to a kanMX selectable marker, allowing further genetic manipulation by synthetic genetic array (SGA)-based, high-throughput methods. We show two such manipulations: barcoding of 440 strains, which enables chemical-genetic suppression analysis, and the construction of arrays of strains carrying different fluorescent markers of subcellular structure, which enables quantitative analysis of phenotypes using high-content screening. Quantitative analysis of a GFP-tubulin marker identified roles for cohesin and condensin genes in spindle disassembly. This mutant collection should facilitate a wide range of systematic studies aimed at understanding the functions of essential genes.

Ämnesord

NATURVETENSKAP  -- Biologi -- Genetik (hsv//swe)

NATURAL SCIENCES  -- Biological Sciences -- Genetics (hsv//eng)

NATURVETENSKAP  -- Biologi -- Biokemi och molekylärbiologi (hsv//swe)

NATURAL SCIENCES  -- Biological Sciences -- Biochemistry and Molecular Biology (hsv//eng)

Nyckelord

Alleles

Databases

Genetic

Genes

Essential

Genes

Fungal

Genes

Lethal

Genetic Engineering

methods

Genetic Loci

Genome

Fungal

Mass Spectrometry

methods

Microarray Analysis

methods

Microscopy

Confocal

Mutation

Phenotype

Plasmids

RNA

Messenger

Saccharomyces cerevisiae

genetics

growth & development

Single-Cell Analysis

Temperature

Tubulin

analysis

Publikations- och innehållstyp

ref (ämneskategori)

art (ämneskategori)