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https://doi.org/10.1007/978-1-...
Part of book or chapter of book . 2011 . Peer-reviewed
License: Springer Nature TDM
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Derivation of Induced Pluripotent Stem Cells by Lentiviral Transduction

Authors: Hubert E, Nethercott; David J, Brick; Philip H, Schwartz;

Derivation of Induced Pluripotent Stem Cells by Lentiviral Transduction

Abstract

This chapter provides a method for reprogramming human dermal fibroblasts into induced pluripotent stem cells (iPSCs) using three lentiviruses containing cDNAs for OCT4 and SOX2, KLF4 and C-MYC, and NANOG and LIN28, respectively. Lentiviral vectors are based on the human immunodeficiency virus (HIV) and provide an effective means for the delivery, integration, and expression of exogenous genes in mammalian cells. Lentiviruses are attractive gene delivery vehicles as they are able to infect both proliferating and nonproliferating cells. Lentiviruses stably integrate into the genome without incurring cellular toxicity and can maintain sustained transgene expression during prolonged host cell proliferation and differentiation. In this protocol, we describe how to prepare lentiviruses, stably transduce human fibroblasts, and identify bona fide iPSC colonies based on morphological similarity to human embryonic stem cell (ESC) colonies and live-cell immunological staining using cell-surface markers of human PSCs such as Tra-1-60 and Tra-1-81.

Related Organizations
Keywords

Staining and Labeling, Cell Survival, Genetic Vectors, Induced Pluripotent Stem Cells, Lentivirus, Cell Culture Techniques, Enzyme-Linked Immunosorbent Assay, Dermis, Fibroblasts, Cellular Reprogramming, Immunohistochemistry, Colony-Forming Units Assay, Kruppel-Like Factor 4, Imaging, Three-Dimensional, Transduction, Genetic, Escherichia coli, Humans, Plasmids

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    20
    popularity
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    Top 10%
    influence
    This indicator reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
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    impulse
    This indicator reflects the initial momentum of an article directly after its publication, based on the underlying citation network.
    Average
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selected citations
These citations are derived from selected sources.
This is an alternative to the "Influence" indicator, which also reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
BIP!Citations provided by BIP!
popularity
This indicator reflects the "current" impact/attention (the "hype") of an article in the research community at large, based on the underlying citation network.
BIP!Popularity provided by BIP!
influence
This indicator reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
BIP!Influence provided by BIP!
impulse
This indicator reflects the initial momentum of an article directly after its publication, based on the underlying citation network.
BIP!Impulse provided by BIP!
20
Top 10%
Top 10%
Average