Functional analysis with a barcoder yeast gene overexpression system.
Systematic analysis of gene overexpression phenotypes provides an insight into gene function, enzyme targets, and biological pathways. Here, we describe a novel functional genomics platform that enables a highly parallel and systematic assessment of overexpression phenotypes in pooled cultures. First, we constructed a genome-level collection of ~5100 yeast barcoder strains, each of which carries a unique barcode, enabling pooled fitness assays with a barcode microarray or sequencing readout. Second, we constructed a yeast open reading frame (ORF) galactose-induced overexpression array by generating a genome-wide set of yeast transformants, each of which carries an individual plasmid-born and sequence-verified ORF derived from the Saccharomyces cerevisiae full-length EXpression-ready (FLEX) collection. We combined these collections genetically using synthetic genetic array methodology, generating ~5100 strains, each of which is barcoded and overexpresses a specific ORF, a set we termed "barFLEX." Additional synthetic genetic array allows the barFLEX collection to be moved into different genetic backgrounds. As a proof-of-principle, we describe the properties of the barFLEX overexpression collection and its application in synthetic dosage lethality studies under different environmental conditions.
Department of Molecular Genetics, University of Toronto, Ontario M5S 1A8, Canada.
Alison C
AC
Douglas
Andrew M
AM
Smith
Sara
S
Sharifpoor
Zhun
Z
Yan
Tanja
T
Durbic
Lawrence E
LE
Heisler
Anna Y
AY
Lee
Owen
O
Ryan
Hendrikje
H
Göttert
Anu
A
Surendra
Dewald
D
van Dyk
Guri
G
Giaever
Charles
C
Boone
Corey
C
Nislow
Brenda J
BJ
Andrews
D019295:Computational Biology / Q000379:methods; D058893:DNA Barcoding, Taxonomic; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D020869:Gene Expression Profiling; D016681:Genome, Fungal; D023281:Genomics / Q000379:methods*; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism
D005656:Fungal Proteins