A pipeline for the generation of shRNA transgenic mice

Dow, L. E., Premsrirut, P. K., Zuber, J., Fellmann, C., McJunkin, K., Miething, C., Park, Y., Dickins, R. A., Hannon, G. J., Lowe, S. W. (2012) A pipeline for the generation of shRNA transgenic mice. Nature Protocols, 7 (2). pp. 374-393. ISSN 17542189 (ISSN)

Abstract

RNA interference (RNAi) is an extremely effective tool for studying gene function in almost all metazoan and eukaryotic model systems. RNAi in mice, through the expression of short hairpin RNAs (shRNAs), offers something not easily achieved with traditional genetic approaches-inducible and reversible gene silencing. However, technical variability associated with the production of shRNA transgenic strains has so far limited their widespread use. Here we describe a pipeline for the generation of miR30-based shRNA transgenic mice that enables efficient and consistent targeting of doxycycline-regulated, fluorescence-linked shRNAs to the Col1a1 locus. Notably, the protocol details crucial steps in the design and testing of miR30-based shRNAs to maximize the potential for developing effective transgenic strains. In all, this 14-week procedure provides a fast and cost-effective way for any laboratory to investigate gene function in vivo in the mouse. © 2012 Nature America, Inc. All rights reserved.

Item Type:	Paper
Uncontrolled Keywords:	doxycycline microRNA miR30 short hairpin RNA unclassified drug animal cell animal experiment article controlled study fluorescence analysis gene function gene locus gene silencing genetic variability genotype in vivo study mouse nonhuman priority journal protein expression protein targeting transgenic mouse Animals Cloning, Molecular Collagen Type I Genetic Engineering Genotyping Techniques Mice Mice, Transgenic MicroRNAs RNA Interference RNA, Small Interfering
Subjects:	bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification bioinformatics > genomics and proteomics > genetics & nucleic acid processing organism description > animal > mammal > rodent > mouse organism description > animal > mammal > rodent bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > shRNA bioinformatics > genomics and proteomics > genetics & nucleic acid processing > transgenic animal
CSHL Authors:	Dow, Lucas E. Fellmann, Christof Hannon, Gregory J. Lowe, Scott W. Miething, Cornelius Premsrirut, Prem K. Zuber, Johannes
Communities:	CSHL Cancer Center Program > Cancer Genetics CSHL Cancer Center Shared Resources > Animal Services CSHL Cancer Center Shared Resources > Bioinformatics Service CSHL Cancer Center Shared Resources > Functional Genomics and Genetics Service CSHL Cancer Center Shared Resources > Gene Targeting Service CSHL labs > Hannon lab CSHL labs > Lowe lab School of Biological Sciences > Publications CSHL Cancer Center Shared Resources > DNA Sequencing Service
Depositing User:	Matt Covey
Date:	2012
Date Deposited:	09 Jan 2013 15:55
Last Modified:	04 Nov 2015 16:06
PMCID:	PMC3724521
Related URLs:	Publisher
URI:	https://repository.cshl.edu/id/eprint/26419

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