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Studying the Replication History of Human B Lymphocytes by Real-Time Quantitative (RQ-)PCR

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Lymphoma

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1956))

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Abstract

The cells of the adaptive immune system, B and T lymphocytes, each generate a unique antigen receptor through V(D)J recombination of their immunoglobulin (Ig) and T-cell receptor (TCR) loci, respectively. Such rearrangements join coding elements to form a coding joint and delete the intervening DNA as circular excision products containing the signal joint. These excision circles are stable structures that cannot replicate and have no function in the cell. Since the coding joint in the genome is replicated with each cell division, the ratio between coding joints and signal joints in a population of B cells can be used as a measure for proliferation. This chapter describes a real-time quantitative (RQ-)PCR-based approach to quantify proliferation through calculating the ratio between coding joints and signal joints of the frequently occurring intronRSS–Kde rearrangements in the IGK light chain locus. Besides its use in normal B-cell biology, quantification of B-cell replication can inform on abnormal proliferation in human diseases and in B-cell neogenesis following depletion therapy.

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Abbreviations

BCR:

B-cell antigen receptor

D:

Diversity

Ig:

Immunoglobulin

IgH:

Ig heavy chain

Igκ:

Ig kappa light chain

Igλ:

Ig lambda light chain

J:

Joining

Kde:

Kappa-deleting element

KREC:

Kappa-deleting recombination excision circle

RAG:

Recombination activating gene

RQ-PCR:

Real-time quantitative PCR

RSS:

Recombination signal sequence

V:

Variable

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Acknowledgments

MCvZ is supported by NHMRC Senior Research Fellowship 1117687.

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Authors and Affiliations

  1. Department of Immunology and Pathology, Central Clinical School, Monash University and The Alfred Hospital, Melbourne, VIC, Australia

    Menno C. van Zelm

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  1. Menno C. van Zelm
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Correspondence to Menno C. van Zelm .

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Editors and Affiliations

  1. Institute of Cell Biology (Cancer Research), Medical School, University of Duisburg-Essen, Nordrhein-Westfalen, Essen, Germany

    Ralf Küppers

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van Zelm, M.C. (2019). Studying the Replication History of Human B Lymphocytes by Real-Time Quantitative (RQ-)PCR. In: Küppers, R. (eds) Lymphoma. Methods in Molecular Biology, vol 1956. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-9151-8_6

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  • DOI: https://doi.org/10.1007/978-1-4939-9151-8_6

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-9150-1

  • Online ISBN: 978-1-4939-9151-8

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