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A rapid and scalable system for studying gene function in mice using conditional RNA interference |
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| Authors: | Premsrirut Prem K Dow Lukas E Kim Sang Yong Camiolo Matthew Malone Colin D Miething Cornelius Scuoppo Claudio Zuber Johannes Dickins Ross A Kogan Scott C Shroyer Kenneth R Sordella Raffaella Hannon Gregory J Lowe Scott W |
| Institution: | 1 Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11724, USA 2 The Watson School of Biological Sciences, Cold Spring Harbor, NY 11724, USA 3 Howard Hughes Medical Institute, Cold Spring Harbor, NY 11724, USA 4 Medical Scientist Training Program, Stony Brook University Medical Center, Stony Brook, New York 11794, USA 5 Department of Pathology, Stony Brook University Medical Center, Stony Brook, New York 11794, USA 6 Molecular Medicine Division, Walter & Eliza Hall Institute of Medical Research, Parkville 3052, Australia 7 Helen Diller Family Comprehensive Cancer Center and Department of Laboratory Medicine, University of California, San Francisco, CA, USA |
| Abstract: | RNA interference is a powerful tool for studying gene function, however, the reproducible generation of RNAi transgenic mice remains a significant limitation. By combining optimized fluorescence-coupled miR30-based shRNAs with high efficiency ES cell targeting, we developed a fast, scalable pipeline for the production of shRNA transgenic mice. Using this system, we generated eight tet-regulated shRNA transgenic lines targeting Firefly and Renilla luciferases, Oct4 and tumor suppressors p53, p16(INK4a), p19(ARF) and APC and demonstrate potent gene silencing and GFP-tracked knockdown in a broad range of tissues in?vivo. Further, using an shRNA targeting APC, we illustrate how this approach can identify predicted phenotypes and also unknown functions for a well-studied gene. In addition, through regulated gene silencing we validate APC/Wnt and p19(ARF) as potential therapeutic targets in T?cell acute lymphoblastic leukemia/lymphoma and lung adenocarcinoma, respectively. This system provides a cost-effective and scalable platform for the production of RNAi transgenic mice targeting any mammalian gene. PAPERCLIP: |
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