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A microarray-based protocol for monitoring the growth of yeast overexpression strains

Nature Protocols volume 1, pages 569–576 (2006)Cite this article

Abstract

Gene overexpression can be used to investigate the biological pathways that are important in the response to a small molecule or other perturbation. To facilitate the use of gene overexpression in the study of small-molecule mechanisms, we developed a microarray-based protocol for monitoring the growth of a pool of yeast strains, each overexpressing a different protein. In this protocol, yeast harboring a set of ∼3,900 galactose-inducible overexpression plasmids are grown in the absence or presence of a small molecule for multiple generations. The plasmids are then extracted from the two populations, processed and labeled in such a manner that their relative concentrations can be determined by competitive hybridization to a microarray. Although this protocol was developed for monitoring a specific set of overexpression plasmids, it could presumably be adapted to monitor yeast that have been transformed with any set of plasmids for which the gene inserts have been spotted, or otherwise arrayed, in a microarray format. This protocol can be completed in approximately 15 hours of hands-on time over the course of several days.

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Figure 1: Outline of protocol.
Figure 2: Typical microarray results.

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Acknowledgements

We acknowledge Chih Long Liu for advice regarding the in vivo transcription amplification strategy. We thank the National Institute of General Medical Sciences (GM38627) for support of this research. S.L.S. is an Investator at the Howard Hughes Medical Institute. R.A.B. was supported by a graduate fellowship from the National Science Foundation.

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  1. Rebecca A Butcher

    Present address: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, 240 Longwood Avenue, Boston, Massachusetts, 02115, USA

Authors and Affiliations

  1. Department of Chemistry and Chemical Biology, Howard Hughes Medical Institute, Harvard University, 12 Oxford Street, Cambridge, 02138, Massachusetts, USA

    Rebecca A Butcher & Stuart L Schreiber

  2. Broad Institute of Harvard and MIT, 7 Cambridge Center, Cambridge, 02142, Massachusetts, USA

    Rebecca A Butcher & Stuart L Schreiber

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  1. Rebecca A Butcher
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  2. Stuart L Schreiber
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Correspondence to Rebecca A Butcher or Stuart L Schreiber.

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Butcher, R., Schreiber, S. A microarray-based protocol for monitoring the growth of yeast overexpression strains. Nat Protoc 1, 569–576 (2006). https://doi.org/10.1038/nprot.2006.80

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