Generation of Human Induced Pluripotent Stem Cells Using Episomal Vectors

Human induced pluripotent stem cells (iPSCs) can be reprogrammed from differentiated somatic cells, such as adult fibroblasts and peripheral blood mononuclear cells, by introducing sets of defined factors. iPSCs closely resemble embryonic stem cells (ESCs) in terms of pluripotency, morphology, proliferative ability, surface markers, expression profiles, and epigenetic status of pluripotent cell-specific genes, among others. In the past decades, the reprogramming has been achieved through various approaches with different induction factors including Sendai virus, adenovirus, episomal DNAs, synthetically modified mRNAs, recombinant proteins, as well as small molecules. Among them, integration-free methods have been promising with a low risk of genomic integration and have the potential for clinical applications. In this chapter, we elaborated a protocol for generating iPSCs from human adult fibroblasts using non-integrative episomal vectors and delineated the strategy of picking iPSC colonies. This protocol basically takes 40 days to generate undifferentiated iPSCs. In summary, this chapter described a method to generate human iPSCs using integration-free approach with episomal vectors and provided a detailed and stepwise procedure for iPSC generation.