International Journal of Biological Sciences

Impact factor
3.873

ISSN 1449-2288

News feeds of IJBS published articles
My Manuscript
My Account

Journal of Biomedicinenew

Theranostics

International Journal of Medical Sciences

Journal of Cancer

Oncomedicine

Journal of Genomics

Journal of Bone and Joint Infection (JBJI)

Nanotheranostics

Journal of Genomics now in PubMed/PubMed Central. Submit manuscript...

PubMed Central Indexed in Journal Impact Factor

Int J Biol Sci 2007; 3(2):91-99. doi:10.7150/ijbs.3.91

Technical Report

RNAi-based conditional gene knockdown in mice using a U6 promoter driven vector

Vivek Shukla, Xavier Coumoul#, Chu-Xia Deng

Genetics of Development and Disease Branch, 10/9N105, National Institute of Diabetes, Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland, MD 20892, USA.
# Present address: INSERM UMR-S 747, Unite de Pharmacologie, Toxicologie et Signalisation Cellulaire, Centre Universitaire des Saints-Pères, 45 rue des Saints-Pères, 75270 Paris Cedex 06, FRANCE

Abstract

RNA interference (RNAi) is a powerful tool widely used for studying gene function in a number of species. We have previously developed an approach that allows conditional expression of a polymerase III promoter based small hairpin RNA (shRNA) in mice using the Cre-LoxP system. This approach uses a U6 promoter, which is inactive due to the presence of a ploxPneo cassette in the promoter; this promoter can be activated after excision of the neo gene in transgenic mice that express a Cre recombinase transgene. As a proof of principle, we have previously knocked down over 95% of Fgfr2 transcripts in mouse germlines, leading to embryonic lethality, while restricting the knockdown to the progress zone of the limb results in live animals with malformation of digits of both the forelimbs and hindlimbs. We now provide a detailed protocol, including a simplified single-step cloning procedure for vector construction. This method provides a fast yet efficient way to decipher gene functions in vivo in a tissue specific manner.

Keywords: RNAi, shRNA, pol III vectors, transgenic mice, protocol

This is an open access article distributed under the terms of the Creative Commons Attribution (CC BY-NC) License. See http://ivyspring.com/terms for full terms and conditions.
How to cite this article:
Shukla V, Coumoul X, Deng CX. RNAi-based conditional gene knockdown in mice using a U6 promoter driven vector. Int J Biol Sci 2007; 3(2):91-99. doi:10.7150/ijbs.3.91. Available from http://www.ijbs.com/v03p0091.htm