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A retroviral strategy that efficiently creates chromosomal deletions in mammalian cells

Nature Methods volume 4, pages 263268 (2007) | Download Citation



Chromosomal deletions, as a genetic tool for functional genomics, remain underexploited for vertebrate stem cells mostly because presently available methods are too labor-intensive. To address this, we developed and validated a set of complementary retroviruses that creates a wide range of nested chromosomal deletions. When applied to mouse embryonic stem cells (ESCs), this retrovirus-based method yielded deletions ranging from 6 kb to 23 Mb (average 2.9 Mb), with an efficiency of 64% for drug-selected clones. Notably, several of the engineered ESC clones, mostly those with large deletions, showed major alteration in cell fate. In comparison to other methods that have also exploited retroviruses for chromosomal engineering, this modified strategy is more efficient and versatile because it bypasses the need for homologous recombination, and thus can be exploited for rapid and extensive functional screens in embryonic and adult stem cells.

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  1. 1.

    , & Nested chromosomal deletions induced with retroviral vectors in mice. Nat. Genet. 24, 92–95 (2000).

  2. 2.

    & Engineering chromosomal rearrangements in mice. Nat. Rev. Genet. 2, 780–790 (2001).

  3. 3.

    , & Efficient Cre-loxP-induced mitotic recombination in mouse embryonic stem cells. Nat. Genet. 30, 66–72 (2002).

  4. 4.

    , , , & Embryonic stem cells and somatic cells differ in mutation frequency and type. Proc. Natl. Acad. Sci. USA 99, 3586–3590 (2002).

  5. 5.

    et al. Trisomy eight in ES cells is a common potential problem in gene targeting and interferes with germ line transmission. Dev. Dyn. 209, 85–91 (1997).

  6. 6.

    & Mouse chromosome engineering for modeling human disease. Annu. Rev. Genomics Hum. Genet. 7, 247–276 (2006).

  7. 7.

    , , & Modeling chromosomes in mouse to explore the function of genes, genomic disorders, and chromosomal organization. PLoS Genet. 2, e86 (2006).

  8. 8.

    et al. Mutagenic insertion and chromosome engineering resource (MICER). Nat. Genet. 36, 867–871 (2004).

  9. 9.

    et al. High-throughput trapping of secretory pathway genes in mouse embryonic stem cells. Nucleic Acids Res. 34, e25 (2006).

  10. 10.

    et al. The International Gene Trap Consortium Website: a portal to all publicly available gene trap cell lines in mouse. Nucleic Acids Res. 34, D642–D648 (2006).

  11. 11.

    et al. Genotype-based screen for ENU-induced mutations in mouse embryonic stem cells. Nat. Genet. 24, 314–317 (2000).

  12. 12.

    , , , & Selection for transgene homozygosity in embryonic stem cells results in extensive loss of heterozygosity. Nat. Genet. 27, 257–258 (2001).

  13. 13.

    , , , & Neonatal lethality and lymphopenia in mice with a homozygous disruption of the c-abl proto-oncogene. Cell 65, 1153–1163 (1991).

  14. 14.

    , & Stable suppression of tumorigenicity by virus-mediated RNA interference. Cancer Cell 2, 243–247 (2002).

  15. 15.

    & Ethanol precipitation of DNA with linear polyacrylamide as carrier. Nucleic Acids Res. 18, 378 (1990).

  16. 16.

    PCR amplification of up to 35-kb DNA with high fidelity and high yield from lambda bacteriophage templates. Proc. Natl. Acad. Sci. USA 91, 2216–2220 (1994).

  17. 17.

    BLAT–the BLAST-like alignment tool. Genome Res. 12, 656–664 (2002).

  18. 18.

    et al. The UCSC Table Browser data retrieval tool. Nucleic Acids Res. 32, D493–D496 (2004).

  19. 19.

    et al. Ensembl 2005. Nucleic Acids Res. 33, D447–D453 (2005).

  20. 20.

    et al. CGHAnalyzer: a stand-alone software package for cancer genome analysis using array-based DNA copy number data. Bioinformatics 21, 3308–3311 (2005).

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We thank P. Chartrand, M. Therrien and colleagues for critically reading the manuscript; V. Paradis, S. Harton and E. Milot from the transgenic facility of IRIC; J. Cowell, M. Rossi and D. McQuaid for the aCGH service (Roswell Park Cancer Institute); J.-P. Laverdure from the bioinformatic service of IRIC;C. Charbonneau from the imaging service of IRIC; N. Fradet, M. Fréchette, A. Fredette, E. St-Hilaire, T. MacRae, C. Rondeau and P. Lussier for technical assistance; A. Nagy for providing the R1 ESCs and the pCX-EYFP construct (Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto); A. Bradley for the pOG231 construct (Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton); R.G. Hawley for the MSCV vectors (The George Washington University Medical Center); M. van Lohuizen for the pRETRO-SUPER construct (The Netherlands Cancer Institute) and R. Jaenisch for the DR-4 mouse strain (Whitehead Institute for Biomedical Research, Massachusetts Institute of Technology). This work was mostly supported by a grant from Génome Québec to G.S. and in part by a grant from the Réseau de Recherche en Transgenèse du Québec to J.H. M.B. is a recipient of a Canadian Institutes of Health Research (CIHR) studentship, and G.S. is a recipient of a Canada Research Chair in molecular genetics of stem cells and a scholar of the Leukemia Lymphoma Society of America.

Author information


  1. Laboratory of Molecular Genetics of Stem Cells, Institute for Research in Immunology and Cancer (IRIC), Université de Montréal, Montréal, Québec, Canada, H2W 1R7.

    • Mélanie Bilodeau
    • , Simon Girard
    •  & Guy Sauvageau
  2. Department of Medicine, Montréal, Québec, Canada, H3C 3J7.

    • Josée Hébert
    •  & Guy Sauvageau
  3. Leukemia Cell Bank of Quebec and Division of Hematology, Maisonneuve-Rosemont Hospital, Montréal, Québec, Canada, H1T 2M2.

    • Josée Hébert
    •  & Guy Sauvageau


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M.B. performed all the experiments and the analyses described herein, except for I-PCR (S.G.), spectral karyotyping and FISH analyses (J.H.). aCGH experiments and chimeras production were conducted by the services mentioned above. M.B. wrote the manuscript, prepared all the figures and performed the experiments under the guidance of G.S.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Guy Sauvageau.

Supplementary information

PDF files

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    Supplementary Fig. 1

    Generation of retroviral vectors.

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    Supplementary Fig. 2

    Cre-induced recombination between integrated proviruses.

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    Supplementary Fig. 3

    Display showing the chromosomal deletions confirmed in ES cells.

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    Supplementary Fig. 4

    Evaluation of inter-chromosomal recombination events.

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    Supplementary Fig. 5

    Full-length gels and blots.

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    Supplementary Table 1

    Summary of the Cre-mediated recombination around 11 randomly chosen loci.

  7. 7.

    Supplementary Table 2

    In vitro differentiation of primary and tertiary clones carrying deletions.

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    Supplementary Methods

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