Abstract
Lentiviral (LV) vectors offer unique advantages over other gene delivery systems, namely the ability to integrate transgenes into the genome of both dividing and nondividing cells. Detailed herein is a simple protocol for the production LV vectors, describing the triple transfection of an LV transfer vector and LV helper plasmids into HEK-293 cells, and the subsequent purification of virions from the cellular media. The current protocol is versatile, and can be easily modified to fit the specific needs of the researcher in order to produce relatively high-titer LV vectors which can be used to transduce a wide variety of cells both in vitro and in vivo.
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References
Knipe DM, Howley PM, Griffin DE (2001) Fundamental virology. Lippincott Williams & Wilkins, Philadelphia. p84
Naldini L et al (1996) In vivo gene delivery and stable transduction of nondividing cells by a lentiviral vector. Science 272:263–267
Naldini L (1998) Lentiviruses as gene transfer agents for delivery to non-dividing cells. Curr Opin Biotechnol 9:457–463
Blomer U et al (1997) Highly efficient and sustained gene transfer in adult neurons with a lentivirus vector. J Virol 71:6641–6649
Akkina RK et al (1996) High-efficiency gene transfer into CD34+ cells with a human immunodeficiency virus type 1-based retroviral vector pseudotyped with vesicular stomatitis virus envelope glycoprotein G. J Virol 70:2581–2585
Kafri T, Blomer U, Peterson DA, Gage FH, Verma IM (1997) Sustained expression of genes delivered directly into liver and muscle by lentiviral vectors. Nat Genet 17:314–317
Case SS et al (1999) Stable transduction of quiescent CD34(+)CD38(−) human hematopoietic cells by HIV-1-based lentiviral vectors. Proc Natl Acad Sci U S A 96:2988–2993
Sutton RE, Wu HT, Rigg R, Böhnlein E, Brown PO (1998) Human immunodeficiency virus type 1 vectors efficiently transduce human hematopoietic stem cells. J Virol 72:5781–5788
Cannon JR, Sew T, Montero L, Burton EA, Greenamyre JT (2011) Pseudotype-dependent lentiviral transduction of astrocytes or neurons in the rat substantia nigra. Exp Neurol 228:41–52
Cronin J, Zhang X-Y, Reiser J (2005) Altering the tropism of lentiviral vectors through pseudotyping. Curr Gene Ther 5:387–398
Semple-Rowland SL, Berry J (2014) Use of lentiviral vectors to deliver and express bicistronic transgenes in developing chicken embryos. Methods 66:466–473. doi:10.1016/j.ymeth.2013.06.026
Geraerts M, Michiels M, Baekelandt V, Debyser Z, Gijsbers R (2005) Upscaling of lentiviral vector production by tangential flow filtration. J Gene Med 7:1299–1310
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This work was supported by the Saint Mary’s Foundation.
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Benskey, M.J., Manfredsson, F.P. (2016). Lentivirus Production and Purification. In: Manfredsson, F. (eds) Gene Therapy for Neurological Disorders. Methods in Molecular Biology, vol 1382. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3271-9_8
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DOI: https://doi.org/10.1007/978-1-4939-3271-9_8
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-3270-2
Online ISBN: 978-1-4939-3271-9
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