A molecular barcoded yeast ORF library enables mode-of-action analysis of bioactive compounds

Cheuk Hei Ho, Leslie Magtanong, Sarah L. Barker, David Gresham, Shinichi Nishimura, Paramasivam Natarajan, Judice L.Y. Koh, Justin Porter, Christopher A. Gray, Raymond J. Andersen, Guri Giaever, Corey Nislow, Brenda Andrews, David Botstein, Todd R. Graham, Minoru Yoshida, Charles Boone

Research output: Contribution to journalArticle

Abstract

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.

Original languageEnglish (US)
Pages (from-to)369-377
Number of pages9
JournalNature Biotechnology
Volume27
Issue number4
DOIs
StatePublished - Apr 2009

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Applied Microbiology and Biotechnology
  • Molecular Medicine
  • Biomedical Engineering

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    Ho, C. H., Magtanong, L., Barker, S. L., Gresham, D., Nishimura, S., Natarajan, P., Koh, J. L. Y., Porter, J., Gray, C. A., Andersen, R. J., Giaever, G., Nislow, C., Andrews, B., Botstein, D., Graham, T. R., Yoshida, M., & Boone, C. (2009). A molecular barcoded yeast ORF library enables mode-of-action analysis of bioactive compounds. Nature Biotechnology, 27(4), 369-377. https://doi.org/10.1038/nbt.1534