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RNAi screening for fat regulatory genes with SRS microscopy

Nature Methods volume 8, pages 135–138 (2011)Cite this article

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Abstract

Identification of genes regulating fat accumulation is important for basic and medical research; genetic screening for those genes in Caenorhabditis elegans, a widely used model organism, requires in vivo quantification of lipids. We demonstrated RNA interference screening based on quantitative imaging of lipids with label-free stimulated Raman scattering (SRS) microscopy, which overcomes major limitations of coherent anti-Stokes Raman scattering microscopy. Our screening yielded eight new genetic regulators of fat storage.

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Figure 1: Visualization of cellular fat storage in lipid droplets using SRS microscopy.
Figure 2: Imaging fat accumulation and distribution in C. elegans by SRS microscopy.
Figure 3: RNAi screening of new fat storage regulatory genes based on in vivo lipid quantification using label-free SRS microscopy.

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Acknowledgements

We thank members of the H. Hutter laboratory (Simon Fraser University) for providing the nre-1(hd20)lin-15b(hd126) strain, members of the C. Sztalryd lab (University of Maryland School of Medicine) for providing perilipin A, ADFP and LSDP5 plasmids, J. Melo and Y. Hao for helping with the membrane receptor RNAi library, A. Soukas and H. Mak for helping with the nuclear hormone receptor RNAi library, Z. Shi for helping with RNAi clone sequencing, X. Zhang and B. Saar for helping on imaging instrumentation, P. Iakova and A. Folick for helping with mammalian cell transfection and C. He and J. Kang for gas chromatography analysis. This work was supported by US National Institutes of Health grants AG034988 (M.C.W.) and EB010244 (X.S.X.) and a predoctoral fellowship from Boehringer Ingelheim Fonds (C.W.F.).

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Author notes

  1. Wei Min

    Present address: Present address: Department of Chemistry, Columbia University, New York, New York, USA.,

  2. Meng C Wang and Wei Min: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Molecular and Human Genetics and Huffington Center on Aging, Baylor College of Medicine, Houston, Texas, USA

    Meng C Wang

  2. Department of Molecular Biology, and Department of Genetics, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, USA

    Meng C Wang & Gary Ruvkun

  3. Department of Chemistry and Chemical Biology, Harvard University, Cambridge, Massachusetts, USA

    Wei Min, Christian W Freudiger & X Sunney Xie

  4. Department of Physics, Harvard University, Cambridge, Massachusetts, USA

    Christian W Freudiger

Authors
  1. Meng C Wang
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  2. Wei Min
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  3. Christian W Freudiger
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  4. Gary Ruvkun
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Contributions

M.C.W., W.M. and X.S.X. conceived the study; M.C.W. and W.M. designed the experiments; M.C.W., W.M. and C.W.F. performed the experiments; M.C.W. analyzed the data; M.C.W., W.M., G.R. and X.S.X. wrote the manuscript.

Corresponding authors

Correspondence to Meng C Wang, Wei Min or X Sunney Xie.

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The authors declare no competing financial interests.

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Supplementary Figures 1– 4 and Supplementary Table 1 (PDF 1583 kb)

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Wang, M., Min, W., Freudiger, C. et al. RNAi screening for fat regulatory genes with SRS microscopy. Nat Methods 8, 135–138 (2011). https://doi.org/10.1038/nmeth.1556

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