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Library Generation and Auxotrophic Selection Assays in Escherichia coli and Thermus thermophilus

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Protein Engineering

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1685))

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Abstract

The selection of optimized enzymes from gene libraries is important, both for basic and applied research. Here, we first describe the generation of plasmid-borne libraries using error-prone PCR and highly competent Escherichia coli cells. We then provide protocols for the use of these libraries for auxotrophic selection assays with E. coli and the extremely thermophilic bacterium Thermus thermophilus as hosts.

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Author information

Authors and Affiliations

  1. Clariant Produkte (Deutschland) GmbH, Semmelweisstrasse 1, 82152, Planegg, Germany

    Jörg Claren

  2. Boehringer Ingelheim Pharma GmbH & Co. KG, 88397, Biberach, Germany

    Thomas Schwab

  3. Institute of Biophysics and Physical Biochemistry, University of Regensburg, Universitätsstrasse 31, 93053, Regensburg, Germany

    Reinhard Sterner

Authors
  1. Jörg Claren
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  2. Thomas Schwab
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  3. Reinhard Sterner
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Corresponding author

Correspondence to Reinhard Sterner .

Editor information

Editors and Affiliations

  1. Department of Biotechnology and Enzyme Catalysis Institute of Biochemistry, Greifswald University, Greifswald, Germany

    Uwe T. Bornscheuer

  2. Protein Biochemistry Institute of Biochemistry, Greifswald University, Greifswald, Germany

    Matthias Höhne

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Claren, J., Schwab, T., Sterner, R. (2018). Library Generation and Auxotrophic Selection Assays in Escherichia coli and Thermus thermophilus . In: Bornscheuer, U., Höhne, M. (eds) Protein Engineering. Methods in Molecular Biology, vol 1685. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7366-8_20

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  • DOI: https://doi.org/10.1007/978-1-4939-7366-8_20

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7364-4

  • Online ISBN: 978-1-4939-7366-8

  • eBook Packages: Springer Protocols

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