High-throughput, quantitative analyses of genetic interactions in E. coli

An array-based high-throughput approach, genetic interaction analysis technology for Escherichia coli (GIANT-coli), now allows comprehensive genetic interaction screens in bacteria. The method uses bacterial conjugation and robotic technology to generate double mutants on a genome-wide scale. In this issue another paper presents eSGA, a very similar approach. 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 .