Efficient derivation and genetic modifications of human pluripotent stem cells on engineered human feeder cell lines

Chunlin Zou, Bin Kuan Chou, Sarah N. Dowey, Kitman Tsang, Xiaosong Huang, Cyndi F. Liu, Cory Smith, Jonathan Yen, Prashant Mali, Yu Alex Zhang, Linzhao Cheng, Zhaohui Ye

Research output: Contribution to journalArticlepeer-review

23 Scopus citations


Derivation of pluripotent stem cells (iPSCs) induced from somatic cell types and the subsequent genetic modifications of disease-specific or patient-specific iPSCs are crucial steps in their applications for disease modeling as well as future cell and gene therapies. Conventional procedures of these processes require co-culture with primary mouse embryonic fibroblasts (MEFs) to support self-renewal and clonal growth of human iPSCs as well as embryonic stem cells (ESCs). However, the variability of MEF quality affects the efficiencies of all these steps. Furthermore, animal sourced feeders may hinder the clinical applications of human stem cells. In order to overcome these hurdles, we established immortalized human feeder cell lines by stably expressing human telomerase reverse transcriptase, Wnt3a, and drug resistance genes in adult mesenchymal stem cells. Here, we show that these immortalized human feeders support efficient derivation of virus-free, integration-free human iPSCs and long-term expansion of human iPSCs and ESCs. Moreover, the drug-resistance feature of these feeders also supports nonviral gene transfer and expression at a high efficiency, mediated by piggyBac DNA transposition. Importantly, these human feeders exhibit superior ability over MEFs in supporting homologous recombination-mediated gene targeting in human iPSCs, allowing us to efficiently target a transgene into the AAVS1 safe harbor locus in recently derived integration-free iPSCs. Our results have great implications in disease modeling and translational applications of human iPSCs, as these engineered human cell lines provide a more efficient tool for genetic modifications and a safer alternative for supporting self-renewal of human iPSCs and ESCs.

Original languageEnglish (US)
Pages (from-to)2298-2311
Number of pages14
JournalStem Cells and Development
Issue number12
StatePublished - Aug 10 2012
Externally publishedYes

ASJC Scopus subject areas

  • Hematology
  • Developmental Biology
  • Cell Biology


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