Generation of Induced Pluripotent Stem Cells with CytoTune, a Non-Integrating Sendai Virus

Primary and Stem Cell Systems, Life Technologies, Carlsbad, CA, USA.
Methods in molecular biology (Clifton, N.J.) (Impact Factor: 1.29). 04/2013; 997:45-56. DOI: 10.1007/978-1-62703-348-0_5
Source: PubMed


One of the major obstacles in generating induced pluripotent stem cells for research or downstream applications is the potential modifications of cellular genome as a result of using integrating viruses during reprogramming. Another major disadvantage of reprogramming cells with integrating vectors is that silencing and activation of transgenes are unpredictable, which may affect terminal differentiation potential and increase the risk of using iPSC-derived cells. Here we describe a protocol for the generation of induced pluripotent stem cells using a non-integrating RNA virus, Sendai virus, to efficiently generate transgene-free iPSCs starting with different cell types as well as in feeder-free conditions.

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Available from: Mohan Vemuri, Jul 14, 2015
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    • "To eliminate the possible effects of insertion and mutagenesis, a lot of modified reprogramming methods have been reported [72], such as non-integration adenoviral102103104105 and sendai viral106107108 vector, episomal vector109110111, minicircle [112], plasmid [113], PiggyBac transposon [114], messenger RNAs [115] and miRNAs [116] and excisable expression systems [117, 118]. Compared with viral methods, nucleic acid based approaches mentioned above are much safer, but these methods still cannot get rid of all the possible risk of tumorigenesis. "
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    • "iPSCs have the capacity to differentiate into patient specific cell types, therefore reducing the likelihood of immunological rejection. Since the first report by Nobel Prize laureate Yamanaka that iPSCs could be generated from fibroblasts through the introduction of specific transcription factors using retroviral systems (Takahashi et al., 2007; Takahashi and Yamanaka, 2006), more effective, non-integrative techniques involving the use of Sendai virus have been described (Lieu et al., 2013). Thus, it is currently possible to generate iPSCs from a person's somatic cells and differentiate them into the required functional cell type (Wilson and Wu, 2015): notably, a first human clinical trial is using iPSC-derived retinal pigment epithelium cells to evaluate their efficacy in treating macular degeneration (Reardon and Cyranoski, 2014). "
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    • "Induced PSC were generated from freshly collected, peripheral blood-derived PBMC (Chou et al., 2011). Cells were transduced with the integration-free CytoTune-iPS Sendai Reprogramming Kit, which utilizes Sendai virus particles of the four Yamanaka factors (Lieu et al., 2013; Takahashi et al., 2007). Transduced cells were plated with MEF feeder cells, and fed iPSC medium with freshly added bFGF (Invitrogen) until small colonies were formed in about 2 weeks. "
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