Dual transposon sequencing profiles the genetic interaction landscape in bacteria
- Justin J. Zik
- Morgan N. Price
- Keisha Hanifa Alma Mayra
- Audrey A. Santoso
- Adam P. Arkin
- Adam M. Deutschbauer
- Lok-To Sham
2025-09-25
Gene redundancy complicates systematic characterization of gene function as single-gene deletions may not produce discernible phenotypes. We report dual transposon sequencing (dual Tn-seq), a platform for assaying the fitness of a comprehensive double mutant pool in parallel. Dual Tn-seq couples random barcode transposon site sequencing with the Cre- lox system, enabling deep sampling of 73% of the 1.3 million possible double gene deletions in Streptococcus pneumoniae . The genetic interactions identified span a wide range of biochemical processes, revealing new factors in presumably well-studied pathways, exemplified by a cytidine triphosphate synthase PyrJ. Moreover, this approach should permit further investigation of growth condition–specific genetic interactions. Because dual Tn-seq does not require the construction of a large array of single mutants, it should be readily adaptable to various microorganisms.