Generation of Lentiviral Vectors for Use in Skeletal Muscle Research

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Generation of Lentiviral Vectors for Use in Skeletal Muscle Research

Authors
  • 1 AIMS , (United States)
Type
Published Article
Journal
Methods in Molecular Biology
Pages
285–295
Identifiers
DOI: 10.1007/978-1-61779-343-1_16
Source
AIMS
License
Green

Abstract

Gene therapy is a promising approach for the treatment of a variety of disorders including genetic diseases and cancer. Among the viral vectors used in gene therapy, the lentiviral vector, based on HIV-1, is the only integrative vector able to transduce nondividing cells. The first generation of lentiviral vector was established in 1996. Since then, other generations of lentiviral vector packaging systems were developed to improve this first vector. In this chapter, we describe these different packaging systems, the generation of lentiviral vector from productive cells, the 293T cell line, and the transduction of myogenic cells with a lentiviral vector as well. The interest and the use of lentiviral vectors started in 1996 when Naldini et al. showed that vectors based on human immunodeficiency virus type-1 (HIV-1) were able to transduce quiescent cells (1). Lentivirus, similarly to retrovirus, is part of the Retroviridae family. Its viral capsid is enveloped and its diameter varies between 80 and 100 nm. It contains two copies of positive-sense single-stranded RNA as well as integrase (IN), protease (PRO), and reverse transcriptase (RT) necessary for its transduction. Lentivirus integration is random but tends to occur in chromosomal regions that are more accessible (2) and more likely in transcriptional active sites (3). The HIV-1 genome is composed of three structural genes: Gag, Pol, and Env. Adding to these structural genes, there are 2 regulatory genes: Tat and Rev, and 4 accessory genes: Vif, Vpr, Vpu, and Nef (4). There are also specific sequences that are present in the lentiviral genome such as long terminal repeats (LTRs),

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