The fitness effect of pairs of mutations is of great interest for the characterization of polygenic complex phenotypes. Current methods rely on the transformation of a host cell with different plasmid-encoded libraries. Though effective, this approach scales quadratically with the number of genes to be tested, making it impractical to survey interactions between many genes. To alleviate this difficulty, we introduce a conjugation-based method that can efficiently generate thousands of gene combinations. In our system, the donor (conjugation proficient) strain is transformed with a genome library constructed in a mobilizable plasmid; the recipient can possess any desired plasmid. The pairwise library is then generated by mixing the two strains together, followed by growth under the selective pressure of interest. We demonstrate the utility of this method by identifying gene interactions that enhance fitness under butanol stress.

Novel conjugation-based method for rapidly screening genetic interactions

245th National Meeting (2013)

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