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High-Titer Production of HIV-Based Lentiviral Vectors in Roller Bottles for Gene and Cell Therapy

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Skin Stem Cells

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1879))

Abstract

Lentiviral vectors are becoming preferred vectors of choice for clinical gene therapy trials due to their safety, efficacy, and the long-term gene expression they provide. Although the efficacy of lentiviral vectors is mainly predetermined by the therapeutic genes they carry, they must be produced at high titers to exert therapeutic benefit for in vivo applications. Thus, there is need for practical, robust, and scalable viral vector production methods applicable to any laboratory setting. Here, we describe a practical lentiviral production technique in roller bottles yielding high-titer third-generation lentiviral vectors useful for in vivo gene transfer applications. CaPO4-mediated transient transfection protocol involving the use of a transfer vector and three different packaging plasmids is employed to generate lentivectors in roller bottles. Following clearance of cellular debris via low-speed centrifugation and filtration, virus is concentrated by high-speed ultracentrifugation over sucrose cushion.

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Acknowledgments

This study is supported by grants from Akdeniz University Scientific Research Administration Division (TYL-2015-1027) and the Scientific and Technological Research Council of Turkey (TUBITAK-112S114).

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Authors and Affiliations

  1. Human Gene and Cell Therapy Center of Akdeniz University Hospitals, Antalya, Turkey

    Hazal Banu Olgun, Hale M. Tasyurek, Ahter Dilsad Sanlioglu & Salih Sanlioglu

Authors
  1. Hazal Banu Olgun
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  2. Hale M. Tasyurek
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  3. Ahter Dilsad Sanlioglu
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  4. Salih Sanlioglu
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Corresponding author

Correspondence to Salih Sanlioglu .

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Editors and Affiliations

  1. Ottawa, ON, Canada

    Kursad Turksen

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Olgun, H.B., Tasyurek, H.M., Sanlioglu, A.D., Sanlioglu, S. (2018). High-Titer Production of HIV-Based Lentiviral Vectors in Roller Bottles for Gene and Cell Therapy. In: Turksen, K. (eds) Skin Stem Cells. Methods in Molecular Biology, vol 1879. Humana Press, New York, NY. https://doi.org/10.1007/7651_2018_150

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  • DOI: https://doi.org/10.1007/7651_2018_150

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-8869-3

  • Online ISBN: 978-1-4939-8870-9

  • eBook Packages: Springer Protocols

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