Derivation of GMP-Compliant Integration-Free hiPSCs Using Modified mRNAs

  • Jens Durruthy Durruthy
  • Vittorio Sebastiano
Part of the Methods in Molecular Biology book series (MIMB, volume 1283)


The clinical use of human induced pluripotent stem cells (hiPSCs) and the development of patients-specific gene and cell therapies rely on the development of fast, reliable, and integration-free methods of derivation of pluripotent stem cells from somatic tissues. Here we describe an integration-free protocol for the rapid derivation of hiPSCs from dermal fibroblasts using modified mRNAs. This method is inexpensive, highly efficient, and makes use of reagents that are xeno-free and chemically defined and can therefore be adopted by any Good Manufacturing Practice (GMP) facility.


Xeno-free hiPSCs GMP Clinical use Modified mRNA Regenerative medicine Cell therapy Gene therapy 


  1. 1.
    Takahashi K, Tanabe K, Ohnuki M, Narita M, Ichisaka T et al (2007) Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell 131:861–872CrossRefPubMedGoogle Scholar
  2. 2.
    Hacein-Bey-Abina S, Von Kalle C, Schmidt M, McCormack MP, Wulffraat N et al (2003) LMO2-associated clonal T cell proliferation in two patients after gene therapy for SCID-X1. Science 302:415–419CrossRefPubMedGoogle Scholar
  3. 3.
    Dowey SN, Huang X, Chou BK, Ye Z, Cheng L (2012) Generation of integration-free human induced pluripotent stem cells from postnatal blood mononuclear cells by plasmid vector expression. Nat Protoc 7:2013–2021CrossRefPubMedCentralPubMedGoogle Scholar
  4. 4.
    Jia F, Wilson KD, Sun N, Gupta DM, Huang M et al (2010) A nonviral minicircle vector for deriving human iPS cells. Nat Methods 7:197–199CrossRefPubMedCentralPubMedGoogle Scholar
  5. 5.
    Cho HJ, Lee CS, Kwon YW, Paek JS, Lee SH et al (2010) Induction of pluripotent stem cells from adult somatic cells by protein-based reprogramming without genetic manipulation. Blood 116:386–395CrossRefPubMedGoogle Scholar
  6. 6.
    Merling RK, Sweeney CL, Choi U, De Ravin SS, Myers TG et al (2013) Transgene-free iPSCs generated from small volume peripheral blood nonmobilized CD34+ cells. Blood 121:e98–e107CrossRefPubMedCentralPubMedGoogle Scholar
  7. 7.
    Yoshida Y, Takahashi K, Okita K, Ichisaka T, Yamanaka S (2009) Hypoxia enhances the generation of induced pluripotent stem cells. Cell Stem Cell 5:237–241CrossRefPubMedGoogle Scholar
  8. 8.
    Durruthy Durruthy J, Ramathal C, Sukhwani M, Fang F, Cui J et al (2014) Fate of induced pluripotent stem cells following transplantation to murine seminiferous tubules. Hum Mol Genet 23:3071–3084CrossRefPubMedCentralPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Jens Durruthy Durruthy
    • 1
    • 2
  • Vittorio Sebastiano
    • 1
    • 2
  1. 1.Institute for Stem Cell Biology and Regenerative Medicine, Department of Obstetrics and GynecologyStanford UniversityStanfordUSA
  2. 2.Institute for Stem Cell Biology and Regenerative Medicine, Department of GeneticsStanford UniversityStanfordUSA

Personalised recommendations