Article abstract
Nature Methods 5, 781 - 787 (2008)
Published online: 1 August 2008 | doi:10.1038/nmeth.1240
High-throughput, quantitative analyses of genetic interactions in E. coli
Athanasios Typas1, Robert J Nichols1, Deborah A Siegele2, Michael Shales3, Sean R Collins3,4, Bentley Lim1, Hannes Braberg3, Natsuko Yamamoto5, Rikiya Takeuchi5, Barry L Wanner6, Hirotada Mori5, Jonathan S Weissman3,4, Nevan J Krogan3 & Carol A Gross1
Abstract
Large-scale genetic interaction studies provide the basis for defining gene function and pathway architecture. Recent advances in the ability to generate double mutants en masse in Saccharomyces cerevisiae have dramatically accelerated the acquisition of genetic interaction information and the biological inferences that follow. Here we describe a method based on F factor–driven conjugation, which allows for high-throughput generation of double mutants in Escherichia coli. This method, termed genetic interaction analysis technology for E. coli (GIANT-coli), permits us to systematically generate and array double-mutant cells on solid media in high-density arrays. We show that colony size provides a robust and quantitative output of cellular fitness and that GIANT-coli can recapitulate known synthetic interactions and identify previously unidentified negative (synthetic sickness or lethality) and positive (suppressive or epistatic) relationships. Finally, we describe a complementary strategy for genome-wide suppressor-mutant identification. Together, these methods permit rapid, large-scale genetic interaction studies in E. coli.
- Department of Microbiology and Immunology, University of California at San Francisco, 600 16th Street, San Francisco, California 94158, USA.
- Department of Biology, Texas A&M University, 3258 TAMU College Station, Texas 77843, USA.
- Department of Cellular and Molecular Pharmacology and The California Institute for Quantitative Biomedical Research, University of California at San Francisco, 1700 4th Street, San Francisco, California 94158, USA.
- Howard Hughes Medical Institute, University of California at San Francisco, 1700 4th Street, San Francisco, California 94158, USA.
- Graduate School of Biological Sciences, Nara Institute of Science and Technology, 8916-5 Takayama, Ikoma, Nara 630-0101, Japan.
- Department of Biological Sciences, Purdue University, LILY 1-228 West Lafayette, Indiana 47907, USA.
Correspondence to: Carol A Gross1 e-mail: cgross@cgl.ucsf.edu
Correspondence to: Nevan J Krogan3 e-mail: krogan@cmp.ucsf.edu
MORE ARTICLES LIKE THIS
These links to content published by NPG are automatically generated.
RESEARCH
Second-harmonic imaging microscopy for visualizing biomolecular arrays in cells, tissues and organismsNature Biotechnology Research (01 Nov 2003)
eSGA: E. coli synthetic genetic array analysisNature Methods Article (01 Sep 2008)
