Abstract
Existing transgenic RNAi resources in Drosophila melanogaster based on long double-stranded hairpin RNAs are powerful tools for functional studies, but they are ineffective in gene knockdown during oogenesis, an important model system for the study of many biological questions. We show that shRNAs, modeled on an endogenous microRNA, are extremely effective at silencing gene expression during oogenesis. We also describe our progress toward building a genome-wide shRNA resource.
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Acknowledgements
The design and construction of the first shRNAs were supported in part by the Janelia Farm Visitor Program. We thank G. Rubin, C. Zuker and T. Laverty for their interest and support; R. Hardy and C. Zuker for the data presented in Supplementary Table 2; B. Haley for helpful discussion on shRNAs; L. Cooley (Yale University) for the gift of the MTD-Gal4 line; and Z. Xuan for help with library design. S. Zusman and M. Tworoger of Genetic Services, Inc. generated the transgenic lines. R.Z. is supported by the Leukemia and Lymphoma Society. B.C. is supported by a PhD fellowship from the Boehringer Ingelheim Fonds. This work was supported by two US National Institute of General Medical Sciences R01 grants (GM067761 and GM084947) to N.P., an EU FP7 European Research Council starting grant to J.B. and contributions from the US National Institute of Neurological Disorders and Stroke.
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J.-Q.N., R.Z. and B.C. carried out major experiments; L.-P.L., L.H., D.Y.-Z., H.-S.S., R.B., M.B. and L.A.P. produced the TRiP lines; P.K. performed the luciferase experiments in ovaries; D.H. and J.B. analyzed the piRNA pathway during oogenesis; and G.J.H. and N.P. supervised the project. R.Z., B.C., J.-Q.N., D.H., J.B., G.J.H. and N.P. wrote the manuscript.
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Supplementary Figures 1–8, Supplementary Tables 1–3 and Supplementary Notes 1–3 (PDF 1347 kb)
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Ni, JQ., Zhou, R., Czech, B. et al. A genome-scale shRNA resource for transgenic RNAi in Drosophila. Nat Methods 8, 405–407 (2011). https://doi.org/10.1038/nmeth.1592
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DOI: https://doi.org/10.1038/nmeth.1592
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