Protocol Update | Published:

Generation of T cell receptor–retrogenic mice: improved retroviral-mediated stem cell gene transfer

Nature Protocols volume 8, pages 18371840 (2013) | Download Citation

Abstract

The use of retrogenic mice offers a rapid and flexible approach to T cell receptor (TCR)-transgenic mice. By transducing bone marrow progenitor cells with a retrovirus that encodes a given TCR-α/β subunit, TCR-retrogenic mice can be generated in as few as 4–6 weeks, whereas conventional TCR transgenics can take 6 months or longer. In this updated protocol, we have increased the efficiency of the bone marrow transduction and bone marrow reconstitution compared with our previously published protocol. The main departure from the previous protocol is the implementation of spin transduction with the viral supernatant instead of coculture with the viral producer cell line. The changes in this protocol improve bone marrow viability, increase consistency of the bone marrow transduction and bone marrow engraftment, and they reduce the ratio of bone marrow donor mice to bone marrow recipients.

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References

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Author information

Affiliations

  1. Department of Immunology, St. Jude Children's Research Hospital, Memphis, Tennessee, USA.

    • Matthew L Bettini
    • , Maria Bettini
    • , Clifford S Guy
    •  & Dario A A Vignali
  2. Barbara Davis Center for Childhood Diabetes, University of Colorado Denver, Aurora, Colorado, USA.

    • Maki Nakayama

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Contributions

M.L.B., M.B., M.N. and C.S.G. contributed experimentally to the improved protocol. D.A.A.V., M.L.B. and M.B. wrote the update.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Dario A A Vignali.

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DOI

https://doi.org/10.1038/nprot.2013.111

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