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Site-Directed Mutagenesis and Its Application in Studying the Interactions of T3S Components
Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR). (Francis)ORCID iD: 0000-0001-6817-9535
Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR). Helmholtz Centre for Infection Research, Braunschweig, Germany. (Francis)
Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR). Department of Biological and Environmental Sciences, Troy University, Troy, AL, USA. (Milton)
Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology). Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR). Institute of Structural and Molecular Biology, University College London and Birkbeck, Malet Street, London, UK. (Francis)
2017 (English)In: Type 3 Secretion Systems: Methods and Protocols / [ed] Matthew L. Nilles and Danielle L. Jessen Condry, Springer-Verlag New York, 2017, 11-31 p.Chapter in book (Refereed)
Abstract [en]

Type III secretion systems are a prolific virulence determinant among Gram-negative bacteria. They are used to paralyze the host cell, which enables bacterial pathogens to establish often fatal infections—unless an effective therapeutic intervention is available. However, as a result of a catastrophic rise in infectious bacteria resistant to conventional antibiotics, these bacteria are again a leading cause of worldwide mortality. Hence, this report describes a pDM4-based site-directed mutagenesis strategy that is assisting in our foremost objective to better understand the fundamental workings of the T3SS, using Yersinia as a model pathogenic bacterium. Examples are given that clearly document how pDM4-mediated site-directed mutagenesis has been used to establish clean point mutations and in-frame deletion mutations that have been instrumental in identifying and understanding the molecular interactions between components of the Yersinia type III secretion system.

Place, publisher, year, edition, pages
Springer-Verlag New York, 2017. 11-31 p.
Series
, Methods in Molecular Biology, ISSN 1064-3745 ; 1531
Keyword [en]
Site-directed mutagenesis, Type III secretion systems, Suicide vector pDM4, Mutant libraries, Genetic-based screens, Protein-protein interaction assays
National Category
Biochemistry and Molecular Biology
Research subject
Microbiology
Identifiers
URN: urn:nbn:se:umu:diva-128091DOI: 10.1007/978-1-4939-6649-3_2PubMedID: 27837478ISBN: 978-1-4939-6647-9ISBN: 978-1-4939-6649-3OAI: oai:DiVA.org:umu-128091DiVA: diva2:1048941
Funder
Swedish Research Council, 2014–2105
Available from: 2016-11-22 Created: 2016-11-22 Last updated: 2016-11-22

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Publisher's full textPubMedhttp://link.springer.com/protocol/10.1007%2F978-1-4939-6649-3_2

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Francis, Matthew S
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Department of Molecular Biology (Faculty of Science and Technology)Umeå Centre for Microbial Research (UCMR)
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