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Site- and time-specific gene targeting in the mouse.
Methods. 2001 May; 24(1):71-80.M

Abstract

The efficient introduction of somatic mutations in a given gene, at a given time, in a specific cell type, will facilitate studies of gene function and the generation of animal models for human diseases. We have established a conditional site-specific recombination system in mice using a new version of the Cre/lox system. The Cre recombinase has been fused to a mutated ligand binding domain of the human estrogen receptor (ER), resulting in a tamoxifen-dependent Cre recombinase, Cre-ER(T), that is activated by tamoxifen, but not by estradiol. Transgenic mice were generated expressing Cre-ER(T) under the control of a cytomegalovirus promoter. Administration of tamoxifen to these transgenic mice induced excision of a chromosomally integrated gene flanked by loxP sites in a number of tissues, whereas no excision could be detected in untreated animals. However, the efficiency of excision varied between tissues, and the highest level (approximately 40%) was obtained in the skin. To determine the efficiency of excision mediated by Cre-ER(T) in a given cell type, Cre-ER(T)-expressing mice were crossed with reporter mice in which expression of Escherichia coli beta-galactosidase can be induced through Cre-mediated recombination. The efficiency and kinetics of this recombination were analyzed at the cellular level in the epidermis of 6- to 8-week-old double transgenic mice. Site-specific excision occurred within a few days of tamoxifen treatment in essentially all epidermis cells expressing Cre-ER(T). These results indicate that cell-specific expression of Cre-ER(T) in transgenic mice can be used for efficient tamoxifen-dependent Cre-mediated recombination at loci containing loxP sites, to generate site-specific somatic mutations in a spatiotemporally controlled manner. This conditional site-specific recombination system should allow the analysis of knockout phenotypes that cannot be addressed by conventional gene targeting.

Authors+Show Affiliations

Institut de Génétique et de Biologie Moléculaire et Cellulaire, CNRS/INSERM/ULP, Collège de France, BP 163 67404 Illkirch Cedex, C.U. de Strasbourg, France.No affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

11327805

Citation

Metzger, D, and P Chambon. "Site- and Time-specific Gene Targeting in the Mouse." Methods (San Diego, Calif.), vol. 24, no. 1, 2001, pp. 71-80.
Metzger D, Chambon P. Site- and time-specific gene targeting in the mouse. Methods. 2001;24(1):71-80.
Metzger, D., & Chambon, P. (2001). Site- and time-specific gene targeting in the mouse. Methods (San Diego, Calif.), 24(1), 71-80.
Metzger D, Chambon P. Site- and Time-specific Gene Targeting in the Mouse. Methods. 2001;24(1):71-80. PubMed PMID: 11327805.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Site- and time-specific gene targeting in the mouse. AU - Metzger,D, AU - Chambon,P, PY - 2001/5/1/pubmed PY - 2001/6/29/medline PY - 2001/5/1/entrez SP - 71 EP - 80 JF - Methods (San Diego, Calif.) JO - Methods VL - 24 IS - 1 N2 - The efficient introduction of somatic mutations in a given gene, at a given time, in a specific cell type, will facilitate studies of gene function and the generation of animal models for human diseases. We have established a conditional site-specific recombination system in mice using a new version of the Cre/lox system. The Cre recombinase has been fused to a mutated ligand binding domain of the human estrogen receptor (ER), resulting in a tamoxifen-dependent Cre recombinase, Cre-ER(T), that is activated by tamoxifen, but not by estradiol. Transgenic mice were generated expressing Cre-ER(T) under the control of a cytomegalovirus promoter. Administration of tamoxifen to these transgenic mice induced excision of a chromosomally integrated gene flanked by loxP sites in a number of tissues, whereas no excision could be detected in untreated animals. However, the efficiency of excision varied between tissues, and the highest level (approximately 40%) was obtained in the skin. To determine the efficiency of excision mediated by Cre-ER(T) in a given cell type, Cre-ER(T)-expressing mice were crossed with reporter mice in which expression of Escherichia coli beta-galactosidase can be induced through Cre-mediated recombination. The efficiency and kinetics of this recombination were analyzed at the cellular level in the epidermis of 6- to 8-week-old double transgenic mice. Site-specific excision occurred within a few days of tamoxifen treatment in essentially all epidermis cells expressing Cre-ER(T). These results indicate that cell-specific expression of Cre-ER(T) in transgenic mice can be used for efficient tamoxifen-dependent Cre-mediated recombination at loci containing loxP sites, to generate site-specific somatic mutations in a spatiotemporally controlled manner. This conditional site-specific recombination system should allow the analysis of knockout phenotypes that cannot be addressed by conventional gene targeting. SN - 1046-2023 UR - https://www.unboundmedicine.com/medline/citation/11327805/Site__and_time_specific_gene_targeting_in_the_mouse_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S1046-2023(01)91159-4 DB - PRIME DP - Unbound Medicine ER -