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T-Cell Repertoire Characterization pp 251–264Cite as

A High-Throughput Strategy for T-Cell Receptor Cloning and Expression

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Part of the Methods in Molecular Biology book series (MIMB,volume 2574)

Abstract

Expression of T-cell receptor (TCR) genes is a critical step for TCR characterization and epitope identification. The recent interest in using specific TCRs for cancer immunotherapy has further increased the demand for practical and robust methods to rapidly clone and express TCRs. We show that a recombination-based cloning protocol facilitates simple and rapid transfer of the TCR transgene into different expression systems. In this protocol, we first constructed all the human TRAV and TRBV genes into individual plasmid. To clone any TCR, we only need to ligate a short CDR3 fragment to its corresponding V gene plasmid using Golden Gate cloning. This strategy significantly improves the efficiency of individual TCR cloning and mutagenesis, providing a flexible high-throughput method for TCR analysis and TCR-mediated therapeutics.

Key words

  • TCR high-throughput cloning
  • Lentivirus package
  • Transfection
  • TCR expression

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  • DOI: 10.1007/978-1-0716-2712-9_12
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Author information

Authors and Affiliations

  1. Institute of Immunity, Transplantation and Infection, Stanford University School of Medicine, Stanford, CA, USA

    Qiong Xia, Huang Huang & Mark M. Davis

  2. Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA, USA

    Mark M. Davis

  3. The Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, CA, USA

    Mark M. Davis

Authors
  1. Qiong Xia
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  2. Huang Huang
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  3. Mark M. Davis
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Corresponding author

Correspondence to Mark M. Davis .

Editor information

Editors and Affiliations

  1. Institute of Immunity, Transplantation and Infection, Stanford University School of Medicine, Stanford, CA, USA

    Huang Huang

  2. Institute of Immunity, Transplantation and Infection, Stanford University School of Medicine, Stanford, CA, USA

    Mark M. Davis

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Xia, Q., Huang, H., Davis, M.M. (2022). A High-Throughput Strategy for T-Cell Receptor Cloning and Expression. In: Huang, H., Davis, M.M. (eds) T-Cell Repertoire Characterization. Methods in Molecular Biology, vol 2574. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2712-9_12

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  • DOI: https://doi.org/10.1007/978-1-0716-2712-9_12

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2711-2

  • Online ISBN: 978-1-0716-2712-9

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