Generation and Characterization of Human Induced Pluripotent Stem Cells via Retroviral Delivery of Yamanaka Factors

Limited access to human samples, especially those from patients with certain genetic defects, was a major challenge for studying disease pathogenesis and regenerative medicine. Over one decade ago, human induced pluripotent stem cells (hiPSCs) were successfully reprogrammed from somatic cells, which are able to either self-renew or differentiate into cell types of all germ layers and thereby serving as an unlimited source for human cells. In this chapter, we described a protocol for the generation and characterization of human iPSCs via retroviral delivery of Yamanaka factors. We first described the culture for starting adult fibroblasts, as well as retrovirus preparation. Later on, we elaborated and discussed the detailed iPSC derivation protocol, under a feeder-free condition, by expressing the Yamanaka factors, that is, the classic four reprogramming factors (OCT4, SOX2, KLF4, and c-MYC). Finally, we delineated the standard assays for the characterization of generated iPSC lines for further use.