title: A set of powerful negative selection systems for unmodified Enterobacteriaceae.
abstract: Creation of defined genetic mutations is a powerful method for dissecting mechanisms of bacterial disease; however, many genetic tools are only developed for laboratory strains. We have designed a modular and general negative selection strategy based on inducible toxins that provides high selection stringency in clinical Escherichia coli and Salmonella isolates. No strain- or species-specific optimization is needed, yet this system achieves better selection stringency than all previously reported negative selection systems usable in unmodified E. coli strains. The high stringency enables use of negative instead of positive selection in phage-mediated generalized transduction and also allows transfer of alleles between arbitrary strains of E. coli without requiring phage. The modular design should also allow further extension to other bacteria. This negative selection system thus overcomes disadvantages of existing systems, enabling definitive genetic experiments in both lab and clinical isolates of E. coli and other Enterobacteriaceae.
authors: National University of Singapore, Department of Medicine, Yong Loo Lin School of Medicine, 1E Kent Ridge Road, NUHS Tower Block, Level 10, Singapore 119074.
authors: Varnica
authors: V
authors: Khetrapal
authors: National University of Singapore, Department of Medicine, Yong Loo Lin School of Medicine, 1E Kent Ridge Road, NUHS Tower Block, Level 10, Singapore 119074.
authors: Kurosh
authors: K
authors: Mehershahi
authors: National University of Singapore, Department of Medicine, Yong Loo Lin School of Medicine, 1E Kent Ridge Road, NUHS Tower Block, Level 10, Singapore 119074.
authors: Shazmina
authors: S
authors: Rafee
authors: National University of Singapore, Department of Medicine, Yong Loo Lin School of Medicine, 1E Kent Ridge Road, NUHS Tower Block, Level 10, Singapore 119074.
authors: Siyi
authors: S
authors: Chen
authors: National University of Singapore, Department of Medicine, Yong Loo Lin School of Medicine, 1E Kent Ridge Road, NUHS Tower Block, Level 10, Singapore 119074.
authors: Chiew Ling
authors: CL
authors: Lim
authors: National University of Singapore, Department of Medicine, Yong Loo Lin School of Medicine, 1E Kent Ridge Road, NUHS Tower Block, Level 10, Singapore 119074 Genome Institute of Singapore, Infectious Diseases Group, 60 Biopolis Street, Genome, #02-01, Singapore 138672 slchen@gis.a-star.edu.sg.
authors: Swaine L
authors: http://orcid.org/0000-0002-0107-2861
authors: SL
authors: Chen
mesh_terms: D000483:Alleles; D001427:Bacterial Toxins / Q000096:biosynthesis / Q000235:genetics; D060846:Cell Engineering; D002876:Chromosomes, Bacterial; D003001:Cloning, Molecular / Q000379:methods; D004755:Enterobacteriaceae / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics* / Q000302:isolation & purification; D056426:Genetic Loci; D006801:Humans; D009154:Mutation; D019779:Salmonella enterica / Q000235:genetics / Q000302:isolation & purification; D014161:Transduction, Genetic
chemical_list: D001427:Bacterial Toxins