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A public genome-scale lentiviral expression library of human ORFs

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Abstract

Functional characterization of the human genome requires tools for systematically modulating gene expression in both loss-of-function and gain-of-function experiments. We describe the production of a sequence-confirmed, clonal collection of over 16,100 human open-reading frames (ORFs) encoded in a versatile Gateway vector system. Using this ORFeome resource, we created a genome-scale expression collection in a lentiviral vector, thereby enabling both targeted experiments and high-throughput screens in diverse cell types.

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Figure 1: Overview of hORFeome V8.1.
Figure 2: Performance of the CCSB-Broad lentiviral expression library.

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Acknowledgements

We thank B. Piqani, I. Budianto, D. Szeto, T. Hirozane-Kishikawa, V. Swearingen, A. MacWilliams, T. Nieland, S. Hoang, J. Bochicchio, S. Young, A. Berlin, C. Russ, M. Garber and members of the Broad Institute Genetic Sequencing Platform who provided technical assistance or advice throughout this project, and J. Zhao, T. Roberts and T. Golub for participation in kinase ORFs subcollection generation. This work was supported by Broad Institute Scientific Planning and Allocation of Resources Committee funding, The Ellison Foundation (D.E.H. and M.V.), Dana-Farber Cancer Institute sponsored research funds to CCSB and Center for Cancer Genome Discovery and US National Institutes of Health R33 CA128625 (W.C.H., D.E.R. and D.E.H.).

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Author notes

  1. Kourosh Salehi-Ashtiani & Haley Hieronymus

    Present address: Present addresses: New York University Abu Dhabi, Abu Dhabi, United Arab Emirates and Center for Genomics and Systems Biology, Department of Biology, New York University, New York, New York, USA (K.S.-A.) and Human Oncology and Pathogenesis Program, Memorial Sloan-Kettering Cancer Center, New York, New York, USA (H.H.).,

  2. Xiaoping Yang, Jesse S Boehm, Xinping Yang, Kourosh Salehi-Ashtiani, Tong Hao, Yun Shen and Rakela Lubonja: These authors contributed equally to this work.

Authors and Affiliations

  1. RNA interference (RNAi) Platform, Broad Institute of Harvard and Massachusetts Institute of Technology (MIT), Cambridge, Massachusetts, USA

    Xiaoping Yang, Rakela Lubonja, Ozan Alkan, Tashfeen Bhimdi, Thomas M Green, Serena J Silver, Cindy Nguyen & David E Root

  2. Cancer Program, Broad Institute of Harvard and MIT, Cambridge, Massachusetts, USA

    Jesse S Boehm, Sapana R Thomas, Cory M Johannessen, Haley Hieronymus & William C Hahn

  3. Center for Cancer Systems Biology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA

    Xinping Yang, Kourosh Salehi-Ashtiani, Tong Hao, Yun Shen, Ryan R Murray, Dawit Balcha, Changyu Fan, Chenwei Lin, Lila Ghamsari, Marc Vidal, William C Hahn & David E Hill

  4. Department of Cancer Biology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA

    Xinping Yang, Kourosh Salehi-Ashtiani, Tong Hao, Yun Shen, Ryan R Murray, Dawit Balcha, Changyu Fan, Chenwei Lin, Lila Ghamsari, Marc Vidal & David E Hill

  5. Department of Genetics, Harvard Medical School, Boston, Massachusetts, USA

    Xinping Yang, Kourosh Salehi-Ashtiani, Tong Hao, Yun Shen, Ryan R Murray, Dawit Balcha, Changyu Fan, Chenwei Lin, Lila Ghamsari, Marc Vidal & David E Hill

  6. Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA

    Cory M Johannessen & William C Hahn

  7. Center for Cancer Genome Discovery, Dana-Farber Cancer Institute, Boston, Massachusetts, USA

    Cory M Johannessen & William C Hahn

  8. Department of Pediatric Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts, USA

    Haley Hieronymus

Authors
  1. Xiaoping Yang
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  2. Jesse S Boehm
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  3. Xinping Yang
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  4. Kourosh Salehi-Ashtiani
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  5. Tong Hao
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  6. Yun Shen
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  7. Rakela Lubonja
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  8. Sapana R Thomas
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  9. Ozan Alkan
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  10. Tashfeen Bhimdi
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  11. Thomas M Green
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  12. Cory M Johannessen
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  13. Serena J Silver
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  14. Cindy Nguyen
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  15. Ryan R Murray
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  16. Haley Hieronymus
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  17. Dawit Balcha
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  18. Changyu Fan
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  19. Chenwei Lin
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  20. Lila Ghamsari
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  21. Marc Vidal
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  22. William C Hahn
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  23. David E Hill
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  24. David E Root
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Contributions

J.S.B., Xia. Y. and D.E.R. wrote the manuscript and, together with D.E.H., K.S.-A. and M.V., designed and supervised the process of creating clonal sequenced ORF collections. Xin. Y., D.B., L.G., J.S.B., S.R.T., H.H., R.R.M., K.S.-A. and C.M.J. generated the starting collections of ORFs. R.L., O.A., J.S.B., C.N., Xia. Y., S.J.S., S.R.T. and C.M.J. created the final libraries, DNA, viruses and pLX vectors, and evaluated expression. T.H., Y.S., C.F., C.L., J.S.B., T.B., T.M.G., Xia. Y. and D.E.R. performed bioinformatic analyses. M.V., W.C.H., D.E.H. and D.E.R. supervised the project.

Corresponding authors

Correspondence to Marc Vidal, William C Hahn, David E Hill or David E Root.

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Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–12, Supplementary Tables 1–4; Supplementary Notes 1–8 (PDF 2703 kb)

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Yang, X., Boehm, J., Yang, X. et al. A public genome-scale lentiviral expression library of human ORFs. Nat Methods 8, 659–661 (2011). https://doi.org/10.1038/nmeth.1638

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  • DOI: https://doi.org/10.1038/nmeth.1638

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