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Activity-Based Screening of Metagenomic Fosmid Libraries for Hydrogen-Uptake Enzymes

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Metagenomics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2555))

Abstract

Here, we outline how to identify hydrogenase enzymes from metagenomic fosmid libraries through an activity-based screening approach. A metagenomic fosmid library is constructed in E. coli and the fosmids are transferred into a hydrogenase deletion mutant of Shewanella oneidensis MR-1 (ΔhyaB) via triparental mating. If a fosmid clone exhibits hydrogen-uptake activity, S. oneidensis’ phenotype is restored and hydrogenase activity is indicated by a color change of the medium from yellow to colorless. The screen enables screening of 48 metagenomic fosmid clones in parallel.

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Acknowledgments

We thank Nicolas Rychlik for constructing the S. oneidensis ΔhyaB mutant.

This work has been supported by research grants DFG PE1549-6/1 and DFG-PE1549-6/3 from the German Research Foundation.

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Authors and Affiliations

  1. Geomicrobiology, Department of Marine Biogeochemistry GEOMAR Helmholtz Centre for Ocean Research Kiel, Wischhofstraße, Kiel, Germany

    Nicole Adam-Beyer & Mirjam Perner

Authors
  1. Nicole Adam-Beyer
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  2. Mirjam Perner
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Corresponding author

Correspondence to Nicole Adam-Beyer .

Editor information

Editors and Affiliations

  1. Department of Microbiology and Biotechnology, University of Hamburg, Hamburg, Germany

    Wolfgang R. Streit

  2. Institut für Mikrobiologie und Genetik, Georg-August-Universität Göttingen, Göttingen, Germany

    Rolf Daniel

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© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

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Adam-Beyer, N., Perner, M. (2023). Activity-Based Screening of Metagenomic Fosmid Libraries for Hydrogen-Uptake Enzymes. In: Streit, W.R., Daniel, R. (eds) Metagenomics. Methods in Molecular Biology, vol 2555. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2795-2_6

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  • DOI: https://doi.org/10.1007/978-1-0716-2795-2_6

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2794-5

  • Online ISBN: 978-1-0716-2795-2

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