TitleA molecular barcoded yeast ORF library enables mode-of-action analysis of bioactive compounds.
Publication TypeJournal Article
Year of Publication2009
AuthorsHo, CHei, Magtanong, L, Barker, SL, Gresham, D, Nishimura, S, Natarajan, P, L Y Koh, J, Porter, J, Gray, CA, Andersen, RJ, Giaever, G, Nislow, C, Andrews, B, Botstein, D, Graham, TR, Yoshida, M, Boone, C
JournalNat Biotechnol
Date Published2009 Apr
KeywordsCloning, Molecular, Gene Library, Genetic Engineering, Open Reading Frames

We present a yeast chemical-genomics approach designed to identify genes that when mutated confer drug resistance, thereby providing insight about the modes of action of compounds. We developed a molecular barcoded yeast open reading frame (MoBY-ORF) library in which each gene, controlled by its native promoter and terminator, is cloned into a centromere-based vector along with two unique oligonucleotide barcodes. The MoBY-ORF resource has numerous genetic and chemical-genetic applications, but here we focus on cloning wild-type versions of mutant drug-resistance genes using a complementation strategy and on simultaneously assaying the fitness of all transformants with barcode microarrays. The complementation cloning was validated by mutation detection using whole-genome yeast tiling microarrays, which identified unique polymorphisms associated with a drug-resistant mutant. We used the MoBY-ORF library to identify the genetic basis of several drug-resistant mutants and in this analysis discovered a new class of sterol-binding compounds.

Alternate JournalNat. Biotechnol.