Author/Authors :
Prem K. Premsrirut، نويسنده , , Lukas E. Dow، نويسنده , , Sang Yong Kim، نويسنده , , Matthew Camiolo، نويسنده , , Colin D. Malone، نويسنده , , Cornelius Miething، نويسنده , , Claudio Scuoppo، نويسنده , , Johannes Zuber، نويسنده , , Ross A. Dickins، نويسنده , , Scott C. Kogan، نويسنده , , Kenneth R. Shroyer، نويسنده , , Raffaella Sordella، نويسنده , , Gregory J. Hannon، نويسنده , , Scott W. Lowe، نويسنده ,
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, p16INK4a, p19ARF 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 p19ARF 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.
