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Rapid and Highly Efficient Method for Scarless Mutagenesis within the Salmonella enterica Chromosome

Direct manipulation of bacterial chromosomes by recombination-based techniques has become increasingly important for both cognitive and applied research. Here we demonstrate, for the first time, the combination of the Red recombinase system with I-SceI endonuclease-based selection of successful reco...

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Detalles Bibliográficos
Autores principales: Blank, Kathrin, Hensel, Michael, Gerlach, Roman G.
Formato: Texto
Lenguaje:English
Publicado: Public Library of Science 2011
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3021506/
https://www.ncbi.nlm.nih.gov/pubmed/21264289
http://dx.doi.org/10.1371/journal.pone.0015763
Descripción
Sumario:Direct manipulation of bacterial chromosomes by recombination-based techniques has become increasingly important for both cognitive and applied research. Here we demonstrate, for the first time, the combination of the Red recombinase system with I-SceI endonuclease-based selection of successful recombinants after electroporation with short synthetic olignucleotides. We show the generation of scarless gene knockouts as well as site-directed mutagenesis using the Salmonella virulence-associated two component signaling system PhoPQ. The presented approach is very versatile for generating in-frame deletions, point mutations or insertions within bacterial chromosomes.