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Ligation-Independent Cloning (LIC) Assembly of TALEN Genes

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Chromosomal Mutagenesis

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1239))

Abstract

Modular DNA binding protein architectures hold the promise of wide application in functional genomic studies. Functionalization of DNA binding proteins, e.g. using the FokI nuclease domain, provides a potent tool to induce DNA double strand breaks at user-defined genomic loci. In this regard, TAL (transcription activator-like) effector proteins, secreted by bacteria of the Xanthomonas family, provide the highest degree of modularity in their DNA binding mode. However, the assembly of large and highly repetitive TALE protein coding genes can be challenging. We describe a ligation-independent cloning (LIC) based method to allow high-throughput assembly of TALE nuclease genes at high fidelity and low effort and cost.

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Acknowledgement

J.L.S.-B. is supported by a scholarship of the Studienstiftung des Deutschen Volkes. V.H. is member of the excellence cluster ImmunoSensation and supported by grants from the German Research Foundation (SFB704 and SFB670) and the European Research Council (ERC‐2009‐StG 243046).

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    Authors and Affiliations

    1. Institute of Molecular Medicine, University Hospital, University of Bonn, 53127, Bonn, Germany

      Jonathan L. Schmid-Burgk, Tobias Schmidt & Veit Hornung

    Authors
    1. Jonathan L. Schmid-Burgk
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    2. Tobias Schmidt
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    3. Veit Hornung
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    Correspondence to Veit Hornung .

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    Editors and Affiliations

    1. Department of Genetics, Genome Engineering and iPSC Center, Washington University School of Medicine, St. Louis, Missouri, USA

      Shondra M. Pruett-Miller

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    Schmid-Burgk, J.L., Schmidt, T., Hornung, V. (2015). Ligation-Independent Cloning (LIC) Assembly of TALEN Genes. In: Pruett-Miller, S. (eds) Chromosomal Mutagenesis. Methods in Molecular Biology, vol 1239. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1862-1_8

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    • DOI: https://doi.org/10.1007/978-1-4939-1862-1_8

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    • Publisher Name: Humana Press, New York, NY

    • Print ISBN: 978-1-4939-1861-4

    • Online ISBN: 978-1-4939-1862-1

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