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Advanced Protocols for Animal Transgenesis pp 477–500Cite as

Induced Pluripotency: Generation of iPS Cells from Mouse Embryonic Fibroblasts

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Abstract

The ability to directly reprogram mammalian adult somatic cells to an undifferentiated pluripotent stage similar to that of embryonic stem cells by introduction of a reduced number of transcription factors has opened new venues in many fields of Biology and Medicine. These reprogrammed cells called iPS cells (induced pluripotent stem cells) represent a powerful tool for the study of cell differentiation and pluripotency and a promise for regenerative therapy. Here, we describe a basic procedure for reprogramming of mouse embryonic fibroblasts (MEFs) to iPS cells by expression of three transcription factors: Oct3/4, Sox2, and Klf4.

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Abbreviations

DMEM:

Dulbecco-Modified Eagles Medium

DMSO:

Dimethyl-Sulfoxide

E12.5:

Embryonic day 12.5. The day in which a vaginal plug is detected is considered day 0.5 of embryonic development

E13.5:

Embryonic day 13.5

EGFP:

Enhanced Green Fluorescent Protein from the jellyfish Aequorea victoria. Enhanced means optimized by mutagenesis for its use in mammalian cells

ES cell:

Embryonic Stem cell

FACS:

Fluorescence Activated Cell Sorting

FCS:

Fetal calf Serum

iPS cell:

Induced Pluripotent Stem cell

KSR:

Knockout-Serum Replacement

LB:

Luria Broth

LTR:

Long Terminal Repeat

MEF:

Mouse Embryonic Fibroblast

N2 :

Nitrogen

PBS:

Phosphate Buffer Saline

PCR:

Polymerase Chain Reaction

Pen/Strep:

Penicillin/Streptomycin

RIPA:

Radio-immuno-precipitation assay

Rpm:

Revolutions per minute

RT:

Room temperature

SCNT:

Somatic Cell Nuclear Transfer

SKY:

Spectral Karyotyping

SSEA-1:

Stage-Specific Mouse Embryonic Antigen-1

WT:

Wild Type

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Acknowledgements

The authors thank Carmen Gómez, Marta Riffo, Jaime Muñoz, Maribel Muñoz and Rosa Serrano for their excellent assistance with tissue culture and mouse work, and Marta Cañamero for the histological analysis of teratomas. We also thank the Animal Facility and Comparative Pathology Units at the CNIO for their valuable technical support.

Author information

Authors and Affiliations

  1. Tumor Suppression Group, Molecular Oncology Program, Spanish National Cancer Research Centre (CNIO), C/Melchor Fernández Almagro 3, 28029, Madrid, Spain

    Han Li & Manuel Serrano

  2. Telomeres and Telomerase Group, Molecular Oncology Program, Spanish National Cancer Research Centre (CNIO), C/Melchor Fernández Almagro 3, 28029, Madrid, Spain

    Katerina Strati & María Blasco

  3. Transgenic Mice Unit, Biotechnology Program, Spanish National Cancer Research Centre (CNIO), C/Melchor Fernández Almagro 3, 28029, Madrid, Spain

    Verónica Domínguez, Javier Martín & Sagrario Ortega

Authors
  1. Han Li
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  2. Katerina Strati
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  3. Verónica Domínguez
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  4. Javier Martín
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  5. María Blasco
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  6. Manuel Serrano
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  7. Sagrario Ortega
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Corresponding author

Correspondence to Sagrario Ortega .

Editor information

Editors and Affiliations

  1. Div. Biology, California Institute of Technology, Pasadena, 91125, California, USA

    Shirley Pease

  2. Transgenic Animal Model Core, University of Michigan, Ann Arbor, 48109-5674, Michigan, USA

    Thomas L. Saunders

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© 2011 Springer-Verlag Berlin Heidelberg

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Li, H. et al. (2011). Induced Pluripotency: Generation of iPS Cells from Mouse Embryonic Fibroblasts. In: Pease, S., Saunders, T. (eds) Advanced Protocols for Animal Transgenesis. Springer Protocols Handbooks. Springer, Berlin, Heidelberg. https://doi.org/10.1007/978-3-642-20792-1_20

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  • DOI: https://doi.org/10.1007/978-3-642-20792-1_20

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  • Publisher Name: Springer, Berlin, Heidelberg

  • Print ISBN: 978-3-642-20791-4

  • Online ISBN: 978-3-642-20792-1

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