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A versatile in vivo system for directed dissection of gene expression patterns

Nature Methods volume 8, pages 231–237 (2011)Cite this article

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Abstract

Tissue-specific gene expression using the upstream activating sequence (UAS)–GAL4 binary system has facilitated genetic dissection of many biological processes in Drosophila melanogaster. Refining GAL4 expression patterns or independently manipulating multiple cell populations using additional binary systems are common experimental goals. To simplify these processes, we developed a convertible genetic platform, the integrase swappable in vivo targeting element (InSITE) system. This approach allows GAL4 to be replaced with any other sequence, placing different genetic effectors under the control of the same regulatory elements. Using InSITE, GAL4 can be replaced with LexA or QF, allowing an expression pattern to be repurposed. GAL4 can also be replaced with GAL80 or split-GAL4 hemi-drivers, allowing intersectional approaches to refine expression patterns. The exchanges occur through efficient in vivo manipulations, making it possible to generate many swaps in parallel. This system is modular, allowing future genetic tools to be easily incorporated into the existing framework.

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Figure 1: The InSITE system.
Figure 2: Molecular and genetic validation of the enhancer-trap exchange.
Figure 3: Functional validation of the QF and LexA enhancer trap swaps.
Figure 4: Functional validation of the split-GAL4 and GAL80 enhancer trap swaps.

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Acknowledgements

We thank members of the Clandinin laboratory for helpful advice; M. Müller (University of Basel), M. Wernet (Stanford University), T. Schwabe (Stanford University), G. Dietzl (Stanford University), J. Bateman (Bowdoin College), L. Luo (Stanford University), C.-H. Lee (US National Institutes of Health) and P. Schedl (Princeton University) for reagents; S. Burns for assistance with experiments; and A. Parks at the Bloomington Drosophila Stock Center. M. Klovstad, L. Luo and T. Schwabe provided valuable comments on the manuscript. M. Spletter helped score antennal lobes. This work was funded by a National Institutes of Health Director's Pioneer Award DP1 OD003530 (T.R.C.) and by National Institutes of Health R01 EY015231 (T.R.C.). D.M.G. and M.A.S. were supported by Stanford Dean's Postdoctoral fellowships.

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Authors and Affiliations

  1. Department of Neurobiology, Stanford University, Stanford, California, USA

    Daryl M Gohl, Marion A Silies, Sheetal Bhalerao, Francisco J Luongo & Thomas R Clandinin

  2. Department of Biological Sciences, Stanford University, Stanford, California, USA

    Xiaojing J Gao

  3. Department of Neuroscience, The Johns Hopkins University School of Medicine, Baltimore, Maryland, USA

    Chun-Chieh Lin & Christopher J Potter

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  1. Daryl M Gohl
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Contributions

D.M.G. and T.R.C. designed the experiments; D.M.G., M.A.S., X.J.G., F.J.L., C.-C.L., C.J.P. and T.R.C. performed the experiments; S.B. provided critical reagents; and D.M.G. and T.R.C. wrote the manuscript.

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Correspondence to Thomas R Clandinin.

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Gohl, D., Silies, M., Gao, X. et al. A versatile in vivo system for directed dissection of gene expression patterns. Nat Methods 8, 231–237 (2011). https://doi.org/10.1038/nmeth.1561

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