Reprogramming of Human Fibroblasts to Induced Pluripotent Stem Cells with Sleeping Beauty Transposon-Based Stable Gene Delivery

Methods in Molecular Biology
Attila Sebe, Zoltán Ivics

Abstract

Human induced pluripotent stem (iPS) cells are a source of patient-specific pluripotent stem cells and resemble human embryonic stem (ES) cells in gene expression profiles, morphology, pluripotency, and in vitro differentiation potential. iPS cells are applied in disease modeling, drug screenings, toxicology screenings, and autologous cell therapy. In this protocol, we describe how to derive human iPS cells from fibroblasts by Sleeping Beauty (SB) transposon-mediated gene transfer of reprogramming factors. First, the components of the non-viral Sleeping Beauty transposon system, namely a transposon vector encoding reprogramming transcription factors and a helper plasmid expressing the SB transposase, are electroporated into human fibroblasts. The reprogramming cassette undergoes transposition from the transfected plasmids into the fibroblast genome, thereby resulting in stable delivery of the reprogramming factors. Reprogramming by using this protocol takes ~4 weeks, after which the iPS cells are isolated and clonally propagated.

Citations

Mar 10, 2016·Stem Cells and Development·Thorsten FriedelIrene C Schneider
Oct 4, 2016·Scientific Reports·Carsten JäckelPeter J Nelson
Aug 18, 2017·Pathology Oncology Research : POR·Ákos GasparicsAttila Sebe
Aug 28, 2020·World Journal of Stem Cells·Dharmendra KumarWilfried A Kues

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