A suite of Gateway® cloning vectors for high-throughput genetic analysis in Saccharomyces cerevisiae

Research output: Contribution to journalResearch articleContributedpeer-review


  • Simon Alberti - , Whitehead Institute for Biomedical Research, Howard Hughes Medical Institute (Author)
  • Aaron D. Gitler - , Whitehead Institute for Biomedical Research (Author)
  • Susan Lindquist - , Whitehead Institute for Biomedical Research, Howard Hughes Medical Institute (Author)


In the post-genomic era, academic and biotechnological research is increasingly shifting its attention from single proteins to the analysis of complex protein networks. This change in experimental design requires the use of simple and experimentally tractable organisms, such as the unicellular eukaryote Saccharomyces cerevisiae, and a range of new high-throughput techniques. The Gateway® system has emerged as a powerful high-throughput cloning method that allows for the in vitro recombination of DNA with high speed, accuracy and reliability. Two Gateway-based libraries of overexpression plasmids containing the entire complement of yeast open reading frames (ORFs) have recently been completed. In order to make use of these powerful resources, we adapted the widely used pRS series of yeast shuttle vectors for use in Gateway-based cloning. The resulting suite of 288 yeast Gateway vectors is based upon the two commonly used GPD and GAL1 promoter expression systems that enable expression of ORFs, either constitutively or under galactose-inducible conditions. In addition, proteins of interest can be fused to a choice of frequently used N- or C-terminal tags, such as EGFP, ECFP, EYFP, Cerulean, monomeric DsRed, HA or TAP. We have made this yeast Gateway® vector kit available to the research community via the non-profit Addgene Plasmid Repository (http://www.addgene.org/ yeast_gateway).


Original languageEnglish
Pages (from-to)913-919
Number of pages7
Issue number10
Publication statusPublished - Oct 2007
Externally publishedYes

External IDs

PubMed 17583893
ORCID /0000-0003-4017-6505/work/161409874



  • Cloning, Gateway®, High-throughput, Vector