Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Automated screening for mutants affecting dopaminergic-neuron specification in C. elegans

Abstract

We describe an automated method to isolate mutant Caenorhabditis elegans that do not appropriately execute cellular differentiation programs. We used a fluorescence-activated sorting mechanism implemented in the COPAS Biosort machine to isolate mutants with subtle alterations in the cellular specificity of GFP expression. This methodology is considerably more efficient than comparable manual screens and enabled us to isolate mutants in which dopamine neurons do not differentiate appropriately.

This is a preview of subscription content

Access options

Buy article

Get time limited or full article access on ReadCube.

$32.00

All prices are NET prices.

Figure 1: Screening for dopaminergic cell-fate mutants.
Figure 2: Relative fluorescence intensity plots between red and green channels of sorted worms.
Figure 3: Phenotypes of isolated dopy mutants.

References

  1. Brenner, S. Genetics 77, 71–94 (1974).

    CAS  PubMed  PubMed Central  Google Scholar 

  2. Jorgensen, E.M. & Mango, S.E. Nat. Rev. Genet. 3, 356–369 (2002).

    CAS  Article  PubMed  Google Scholar 

  3. Chalfie, M., Tu, Y., Euskirchen, G., Ward, W.W. & Prasher, D.C. Science 263, 802–805 (1994).

    CAS  Article  PubMed  Google Scholar 

  4. Pulak, R. Methods Mol. Biol. 351, 275–286 (2006).

    PubMed  Google Scholar 

  5. Abeliovich, A. & Hammond, R. Dev. Biol. 304, 447–454 (2007).

    CAS  Article  PubMed  Google Scholar 

  6. Nass, R. et al. J. Neurochem. 94, 774–785 (2005).

    CAS  Article  PubMed  Google Scholar 

  7. Nass, R., Hall, D.H., Miller, D.M., III & Blakely, R.D. Proc. Natl. Acad. Sci. USA 99, 3264–3269 (2002).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  8. Chase, D.L., Pepper, J.S. & Koelle, M.R. Nat. Neurosci. 7, 1096–1103 (2004).

    CAS  Article  PubMed  Google Scholar 

  9. Davis, M.W. et al. BMC Genomics 6, 118 (2005).

    Article  PubMed  PubMed Central  Google Scholar 

  10. Zhao, C. & Emmons, S.W. Nature 373, 74–78 (1995).

    CAS  Article  PubMed  Google Scholar 

  11. Hack, M.A. et al. Nat. Neurosci. 8, 865–872 (2005).

    CAS  Article  PubMed  Google Scholar 

  12. Guenther, C. & Garriga, G. Development 122, 3509–3518 (1996).

    CAS  PubMed  Google Scholar 

  13. Chung, K., Crane, M.M. & Lu, H. Nat. Methods 5, 637–643 (2008).

    CAS  Article  PubMed  Google Scholar 

  14. Boulin, T. & Bessereau, J.L. Nat. Protoc. 2, 1276–1287 (2007).

    CAS  Article  PubMed  Google Scholar 

  15. Sarin, S., Prabhu, S., O'Meara, M.M., Pe'er, I. & Hobert, O. Nat. Methods advance online publication 1 August 2008 (doi:10.1038/nmeth.1249).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We thank A. Kruyer, A. Pretorian, J. Recio and Q. Chen for technical assistance, S. Sarin for calculating the degree of saturation, V. Bertrand and F. Zhang for communicating unpublished results, and S. Mitani (Tokyo Women's Medical University School of Medicine) for providing lin-32 deletion alleles. This work was funded by the US National Institutes of Health (R01NS039996-05; R01NS050266-03), the Howard Hughes Medical Institute, an European Molecular Biology Organization long-term fellowship to M.D. and N.F., and a Marie Curie Outgoing International fellowship to N.F.

Author information

Authors and Affiliations

Authors

Contributions

M.D. conducted the genetic screens (manual and sorter), implemented the sorter strategy, mapped and characterized mutants, and co-wrote the paper; N.F. implemented the sorter strategy and conducted sorter screens; A.C.L. conducted manual screens; A.B. assisted in mapping mutants; and O.H. initiated and supervised the project, and co-wrote the paper.

Corresponding authors

Correspondence to Maria Doitsidou or Oliver Hobert.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–2, Supplementary Tables 1–6, Supplementary Methods (PDF 3378 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Doitsidou, M., Flames, N., Lee, A. et al. Automated screening for mutants affecting dopaminergic-neuron specification in C. elegans. Nat Methods 5, 869–872 (2008). https://doi.org/10.1038/nmeth.1250

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nmeth.1250

Further reading

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing