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Defeating Randomness – Targeted Integration as a Boost for Biotechnology

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Cell Line Development

Part of the book series: Cell Engineering ((CEEN,volume 6))

Abstract

Genetic modification of mammalian cells is a prerequisite for the production of recombinant proteins, virus like particles, and viruses. For most applications long-term stability of such modifications is needed. Apart from episomal approaches which have not yet been sufficiently explored, integration of transgenes into the chromosomal DNA of the host cell is regarded to be the method of choice. Due to strong influences of the chromosomal surroundings on the expression of transgenes, targeted integration has obvious advantages over classical random integration procedures. Site directed integration leads to predictable expression properties, circumvents screening, is fast and provides high safety and is therefore advantageous for the integration of transgenes. In this chapter recombinase mediated cassette exchange (RMCE) using heterologous recombinases is described as an efficient and reliable method to target (integrate and replace) transgenes by site directed engineering of defined chromosomal sites for recombinant protein and virus expression. This technique, however, requires that the site of interest has been tagged before by specific sequence motifs. Thus, the chapter highlights current and forthcoming methods to (find and) tag chromosomal sites in order to make RMCE possible. This includes screening of randomly integrated reporters and spontaneous or Zinc Finger Nuclease enhanced homologous recombination.

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Authors and Affiliations

  1. Department of Gene Regulation and Differentiation and Research Group Model Systems for Infection and Immunity, Helmholtz Centre for Infection Research, Inhoffenstrasse 7, D-38124, Braunschweig, Germany

    L. Gama-Norton, P. Riemer, U. Sandhu, K. Nehlsen, R. Schucht, H. Hauser & D. Wirth

  2. Instituto de Biologia Experimental e Tecnológica, Universidade Nova de Lisboa (IBET/ITQB-UNL), Animal Cell Technology, Oeiras, Portugal

    L. Gama-Norton

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  1. L. Gama-Norton
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  2. P. Riemer
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  3. U. Sandhu
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  4. K. Nehlsen
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  5. R. Schucht
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  6. H. Hauser
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  7. D. Wirth
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Corresponding author

Correspondence to H. Hauser .

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Editors and Affiliations

  1. Dept. Chemical & Biochemical, University College, Dublin, Belfield, Dublin 4, Ireland

    Mohamed Al-Rubeai

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Gama-Norton, L. et al. (2009). Defeating Randomness – Targeted Integration as a Boost for Biotechnology. In: Al-Rubeai, M. (eds) Cell Line Development. Cell Engineering, vol 6. Springer, Dordrecht. https://doi.org/10.1007/978-90-481-2245-5_3

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