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Generating Chromosome-Located Transcriptional Fusions to Fluorescent Proteins for Single-Cell Gene Expression Analysis in Pseudomonas syringae

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Host-Pathogen Interactions

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1734))

Abstract

The last decade has seen significant effort directed toward the role of phenotypic heterogeneity in bacterial adaptation. Phenotypic heterogeneity usually refers to phenotypic diversity that takes place through nongenetic means, independently of environmental induced variation. Recent findings are changing how microbiologists analyze bacterial behavior, with a shift from traditional assays averaging large populations to single-cell analysis focusing on bacterial individual behavior. Fluorescence-based methods are often used to analyze single-cell gene expression by flow cytometry, fluorescence microscopy and/or microfluidics. Moreover, fluorescence reporters can also be used to establish where and when are the genes of interest expressed. In this chapter, we use the model bacterial plant pathogen Pseudomonas syringae to illustrate a method to generate chromosome-located transcriptional gene fusions to fluorescent reporter genes, without affecting the function of the gene of interest.

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Acknowledgments

The work was supported by a Project Grant from Ministerio de Economía y Competitividad (Spain) (BIO2015-64391-R) awarded to C.R. Beuzón and J. Ruiz-Albert. M.G. and J.R. acknowledge support from the BBSRC/EPSRC Synthetic Biology Research Centre (BB/M017982/1) funded under the UK Research Councils’ Synthetic Biology for Growth Programme. The work was cofunded by Fondos Europeos de Desarrollo Regional (FEDER).

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Authors and Affiliations

  1. Dpto. Biología Celular, Genética y Fisiología, Instituto de Hortofruticultura Subtropical y Mediterránea, Universidad de Málaga-Consejo Superior de Investigaciones Científicas (IHSM-UMA-CSIC), Málaga, Spain

    José S. Rufián, Diego López-Márquez, Nieves López-Pagán, Javier Ruiz-Albert & Carmen R. Beuzón

  2. School of Life Sciences, University of Warwick, Conventry, UK

    Murray Grant

Authors
  1. José S. Rufián
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  2. Diego López-Márquez
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  3. Nieves López-Pagán
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  4. Murray Grant
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  5. Javier Ruiz-Albert
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  6. Carmen R. Beuzón
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Corresponding author

Correspondence to Carmen R. Beuzón .

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Editors and Affiliations

  1. Departamento de Biología Molecular e Ingeniería Bioquímica, Centro Andaluz de Biología del Desarrollo, CSIC/Universidad Pablo de, Olavide/Junta de Andalucía, Seville, Spain

    Carlos Medina

  2. Department of Microbiology, University of Seville, Sevilla, Sevilla, Spain

    Francisco Javier López-Baena

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Rufián, J.S., López-Márquez, D., López-Pagán, N., Grant, M., Ruiz-Albert, J., Beuzón, C.R. (2018). Generating Chromosome-Located Transcriptional Fusions to Fluorescent Proteins for Single-Cell Gene Expression Analysis in Pseudomonas syringae . In: Medina, C., López-Baena, F. (eds) Host-Pathogen Interactions. Methods in Molecular Biology, vol 1734. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7604-1_15

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  • DOI: https://doi.org/10.1007/978-1-4939-7604-1_15

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7603-4

  • Online ISBN: 978-1-4939-7604-1

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