A rapid and scalable system for studying gene function in mice using conditional RNA interference.

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:

• Authors: Prem K Premsrirut, Lukas E Dow, Sang Yong Kim, Matthew Camiolo, Colin D Malone, Cornelius Miething, Claudio Scuoppo, Johannes Zuber, Ross A Dickins, Scott C Kogan, Kenneth R Shroyer, Raffaella Sordella, Gregory J Hannon, Scott W Lowe
• Journal: Cell (Cell) Vol. 145 Issue 1 Pg. 145-58 (Apr 01 2011) ISSN: 1097-4172 [Electronic] United States
• PMID: 21458673 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)
• Copyright: Copyright © 2011 Elsevier Inc. All rights reserved.

Chemical References

• MicroRNAs
• RNA, Small Interfering
• Wnt Proteins

Topics

• Adenocarcinoma (genetics, therapy)
• Animals
• Embryonic Stem Cells (metabolism)
• Gene Knockdown Techniques (economics, methods)
• Lung Neoplasms (genetics, therapy)
• Mice
• Mice, Transgenic
• MicroRNAs (genetics)
• Precursor T-Cell Lymphoblastic Leukemia-Lymphoma (genetics, therapy)
• RNA Interference
• RNA Processing, Post-Transcriptional
• RNA, Small Interfering (genetics)
• Signal Transduction
• Wnt Proteins (metabolism)