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Random and targeted transgene insertion in Caenorhabditis elegans using a modified Mos1 transposon

Nature Methods volume 11, pages 529534 (2014) | Download Citation

Abstract

We have generated a recombinant Mos1 transposon that can insert up to 45-kb transgenes into the Caenorhabditis elegans genome. The minimal Mos1 transposon (miniMos) is 550 bp long and inserts DNA into the genome at high frequency (60% of injected animals). Genetic and antibiotic markers can be used for selection, and the transposon is active in C. elegans isolates and Caenorhabditis briggsae. We used the miniMos transposon to generate six universal Mos1-mediated single-copy insertion (mosSCI) landing sites that allow targeted transgene insertion with a single targeting vector into permissive expression sites on all autosomes. We also generated two collections of strains: a set of bright fluorescent insertions that are useful as dominant, genetic balancers and a set of lacO insertions to track genome position.

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Acknowledgements

We thank B. Waterston (University of Washington), A. Sapir and P. Sternberg (California Institute of Technology), and the NemaGENETAG consortium for strains; B. Meyer (UC Berkeley) and P. Meister (University of Bern) for validating lacO insertions; the J. Chin (MRC, University of Cambridge), D. Dupuy (University of Bordeaux), B. Lehner (EMBL-CRG, Systems Biology Unit, Barcelona) and G. Seydoux (John Hopkins University) labs for plasmids; M. Maduro (UC Riverside) for improving mosSCI insertion frequency; and K. Hoe for expert technical assistance. Some strains were provided by the Caenorhabditis Genetics Center (CGC), which is funded by US National Institutes of Health (NIH) Office of Research Infrastructure Programs (P40 OD010440). This work was supported by the Carlsberg Foundation (C.F.-J.), NIH grant 1R01GM095817 (E.M.J.), US National Science Foundation grant NSF IOS-0920069 (E.M.J.) and the Howard Hughes Medical Institute (E.M.J.). The Mosmid engineering work was supported by the Max Planck Society (MPG) Initiative “BAC TransgeneOmics” and the NIH ModENCODE project. Work in the laboratory of D.G.M. was supported by the Canadian Institute for Health Research. Work in the laboratory of D.G.M. was supported by the Canadian Institute for Health Research and the Canadian Institute for Advanced Research.

Author information

Affiliations

  1. Howard Hughes Medical Institute, University of Utah, Salt Lake City, Utah, USA.

    • Christian Frøkjær-Jensen
    • , M Wayne Davis
    •  & Erik M Jorgensen
  2. Department of Biology, University of Utah, Salt Lake City, Utah, USA.

    • Christian Frøkjær-Jensen
    • , M Wayne Davis
    • , Matthew LaBella
    •  & Erik M Jorgensen
  3. Danish National Research Foundation Centre for Cardiac Arrhythmia, University of Copenhagen, Copenhagen, Denmark.

    • Christian Frøkjær-Jensen
  4. TransgeneOmics, Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany.

    • Mihail Sarov
    •  & Andrei Pozniakovsky
  5. Department of Zoology, University of British Columbia, Vancouver, British Columbia, Canada.

    • Jon Taylor
    • , Stephane Flibotte
    •  & Donald G Moerman

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Contributions

C.F.-J. designed experiments under the supervision of E.M.J. and M.W.D. C.F.-J., M.S., A.P., J.T., M.L. and S.F. performed the research. C.F.-J. performed molecular biology, injections, imaging and genetics; M.L. generated mapping strains; M.S. and A.P. performed fosmid recombineering; and J.T., S.F. and D.G.M. performed comparative genome hybridization. C.F.-J. and E.M.J. wrote the paper with input from all coauthors.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Christian Frøkjær-Jensen or Erik M Jorgensen.

Supplementary information

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    Supplementary Text and Figures

    Supplementary Figures 1–9, Supplementary Note and Supplementary Protocols

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    Supplementary Table 1

    Strains, Plasmids, Oligos and Universal insertion sites

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DOI

https://doi.org/10.1038/nmeth.2889

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