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A lentivirus-based system to functionally silence genes in primary mammalian cells, stem cells and transgenic mice by RNA interference

Nature Genetics volume 33, pages 401–406 (2003)Cite this article

A Corrigendum to this article was published on 01 June 2007

A Corrigendum to this article was published on 01 June 2003

This article has been updated

Abstract

RNA interference (RNAi) has recently emerged as a specific and efficient method to silence gene expression in mammalian cells either by transfection of short interfering RNAs (siRNAs; ref. 1) or, more recently, by transcription of short hairpin RNAs (shRNAs) from expression vectors and retroviruses2,3,4,5,6,7,8,9,10. But the resistance of important cell types to transduction by these approaches, both in vitro and in vivo11, has limited the use of RNAi. Here we describe a lentiviral system for delivery of shRNAs into cycling and non-cycling mammalian cells, stem cells, zygotes and their differentiated progeny. We show that lentivirus-delivered shRNAs are capable of specific, highly stable and functional silencing of gene expression in a variety of cell types and also in transgenic mice. Our lentiviral vectors should permit rapid and efficient analysis of gene function in primary human and animal cells and tissues and generation of animals that show reduced expression of specific genes. They may also provide new approaches for gene therapy.

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Figure 1: Stable gene silencing and production of processed shRNAs in a T-cell line by a lentiviral vector.
Figure 2: Functional silencing of genes in cycling and non-cycling primary immune cells in vitro and in vivo by a lentiviral vector.
Figure 3: Functional silencing of genes in ES cell–derived mice by a lentiviral vector.
Figure 4: Functional silencing of genes in transgenic mice derived from lentivirus-injected zygotes.

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  • 14 May 2007

    The authors wish to remove panels b, d, and e of Figure 2 because they were falsified or fabricated. Figure 2 shows the functional silencing of genes in primary cells by lentiviral-mediated RNAi. The authors showed that primary activated T cells can be successfully transduced by lentivirus and that RNAi-targeted surface markers are downregulated in transduced cells (Fig. 2a). This knockdown leads to a deficiency in T cell proliferation in response to IL-2 (Fig. 2c). These conclusions remain intact. Figure 2b demonstrates the transduction of bone marrow–derived dendritic cells and the knockdown of bim expression. Figure 2d shows the transduction of hematopoietic stem cells (HSCs) and the expression of GFP marker in splenocytes from chimeras derived from transduced HSCs. Figure 2e shows the reduction in CD8+ cells harvested in CD8-knockdown bone marrow chimeras. However, the data supporting the transduction of dendritic cells or HSCs with the lentiviral vector pLL3.7 and the resulting knockdown has been determined to be falsified or fabricated, invalidating the conclusions drawn from these panels (Fig. 2b,d,e). We have no reason to believe that the other results and interpretations in this paper need to be corrected. This correction follows an investigation by the Massachusetts Institute of Technology into scientific misconduct by Luk Van Parijs, the communicating author of the paper. The investigation also found that Dr. Van Parijs was solely responsible for the scientific misconduct that resulted in the falsified or fabricated data in this paper. In addition, the authors wish to revise a Corrigendum regarding this paper that was published at Dr. Van Parijs' request (Nat. Genet. 34, 231; 2003). The Corrigendum contained a revised author list that mistakenly omitted Mingdi Zhang, an author on the original paper. The complete author list is as follows: Douglas A. Rubinson, Christopher P. Dillon, Adam V. Kwiatkowski, Claudia Sievers, Lili Yang, Johnny Kopinja, Dina L. Rooney, Mingdi Zhang, Melanie M. Ihrig, Michael T. McManus, Frank B. Gertler, Martin L. Scott & Luk Van Parijs.That Corrigendum also acknowledged partial support from the Broad Institute for the work reported in the paper. The Broad Institute in fact did not support any of that work, and the acknowledgment of its support was mistaken.Editor’s note: all of the authors, with the exception of Luk Van Parijs, agree with the wording of this Corrigendum.

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Acknowledgements

We would like to thank J. Bear, J. Chen, P. Sharp, C. Lois and D. Baltimore for their advice and D. Chojnacky, A. Antov, K. Layer, B. Haines, G. Paradis and the Flow Cytometry facility for their technical support. This study was supported by a grant from the David Koch Cancer Research Fund, a Career Development Award from the Arthritis Foundation and the Juvenile Diabetes Foundation (to L.V.P.), a grant from the US National Institutes of Health and a WM Keck Distinguished Young Scholar Award (to F.B.G.). D.A.R. is supported by the Medical Scientist Training Program. C.P.D. is a Howard Hughes Medical Institute Predoctoral Fellow. M.T.M. is a fellow of the Cancer Research Institute. A.V.K. is supported by an Anna Fuller Predoctoral Scholarship.

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Author notes

  1. Douglas A Rubinson and Christopher P Dillon: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Biology, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, 02139, Massachusetts, USA

    Douglas A Rubinson, Christopher P Dillon, Adam V Kwiatkowski, Claudia Sievers, Michael T McManus, Frank B Gertler & Luk Van Parijs

  2. Center for Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA

    Christopher P Dillon, Claudia Sievers, Michael T McManus & Luk Van Parijs

  3. Free University Berlin, Institute of Biochemistry, Berlin, Germany

    Claudia Sievers

  4. California Institute of Technology, Pasadena, California, USA

    Lili Yang

  5. Biogen Inc., Cambridge, Massachusetts, USA

    Johnny Kopinja, Mingdi Zhang & Martin L Scott

  6. Division of Comparative Medicine, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, 02139, Massachusetts, USA

    Dina L Rooney & Melanie M Ihrig

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Correspondence to Luk Van Parijs.

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Rubinson, D., Dillon, C., Kwiatkowski, A. et al. A lentivirus-based system to functionally silence genes in primary mammalian cells, stem cells and transgenic mice by RNA interference. Nat Genet 33, 401–406 (2003). https://doi.org/10.1038/ng1117

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