Genome editing of human embryonic stem cells and induced pluripotent stem cells with zinc finger nucleases for cellular imaging

Yongming Wang, Wendy Y. Zhang, Shijun Hu, Feng Lan, Andrew S. Lee, Bruno Huber, Leszek Lisowski, Ping Liang, Mei Huang, Patricia E. De Almeida, Jong H. Won, Ning Sun, Robert C. Robbins, Mark A. Kay, Fyodor D. Urnov, Joseph C. Wu

Research output: Contribution to journalArticlepeer-review

78 Scopus citations


RATIONALE: Molecular imaging has proven to be a vital tool in the characterization of stem cell behavior in vivo. However, the integration of reporter genes has typically relied on random integration, a method that is associated with unwanted insertional mutagenesis and positional effects on transgene expression. OBJECTIVE: To address this barrier, we used genome editing with zinc finger nuclease (ZFN) technology to integrate reporter genes into a safe harbor gene locus (PPP1R12C, also known as AAVS1) in the genome of human embryonic stem cells and human induced pluripotent stem cells for molecular imaging. METHODS AND RESULTS: We used ZFN technology to integrate a construct containing monomeric red fluorescent protein, firefly luciferase, and herpes simplex virus thymidine kinase reporter genes driven by a constitutive ubiquitin promoter into a safe harbor locus for fluorescence imaging, bioluminescence imaging, and positron emission tomography imaging, respectively. High efficiency of ZFN-mediated targeted integration was achieved in both human embryonic stem cells and induced pluripotent stem cells. ZFN-edited cells maintained both pluripotency and long-term reporter gene expression. Functionally, we successfully tracked the survival of ZFN-edited human embryonic stem cells and their differentiated cardiomyocytes and endothelial cells in murine models, demonstrating the use of ZFN-edited cells for preclinical studies in regenerative medicine. CONCLUSION: Our study demonstrates a novel application of ZFN technology to the targeted genetic engineering of human pluripotent stem cells and their progeny for molecular imaging in vitro and in vivo.

Original languageEnglish (US)
Pages (from-to)1494-1503
Number of pages10
JournalCirculation research
Issue number12
StatePublished - Dec 2012
Externally publishedYes


  • homologous recombination
  • induced pluripotent stem cells
  • molecular imaging
  • reporter gene
  • zinc finger nuclease

ASJC Scopus subject areas

  • Physiology
  • Cardiology and Cardiovascular Medicine


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