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Building a zoo of mice for genetic analyses: A comprehensive protocol for the rapid generation of BAC transgenic mice


Johansson, T; Broll, I; Frenz, T; Hemmers, S; Becher, B; Zeilhofer, H U; Buch, T (2010). Building a zoo of mice for genetic analyses: A comprehensive protocol for the rapid generation of BAC transgenic mice. Genesis: The Journal of Genetics and Development, 48(4):264-280.

Abstract

Transgenic mice are highly valuable tools for biological research as they allow cell type-specific expression of functionally instrumental genes. In this protocol, the generation of bacterial artificial chromosome (BAC) transgenic constructs is described. We give an overview of different transgenic inserts, such as fluorescent proteins (alone or in combination with Cre variants), diphtheria toxin receptor, lacZ, and light-activated ion channels. The most reliable and versatile approach to express these genes is by using BACs, which allow "highjacking" of the expression pattern of a gene without characterizing its transcriptional control elements. Here, we describe the necessary cloning techniques compared with conventional transgenesis. With the provided "toolbox" of already available transgene constructs, the generation of the BAC transgenes is made easy and rapid. We provide a comprehensive outline how to insert the different transgenes into a chosen BAC by either ET cloning or recombineering. We also describe in detail the methods to identify the correct insertion and the integrity of the final BAC construct, and finally, the preparation of the BAC DNA for oocyte injection is described. genesis 00:1-17, 2010. (c) 2010 Wiley-Liss, Inc.

Transgenic mice are highly valuable tools for biological research as they allow cell type-specific expression of functionally instrumental genes. In this protocol, the generation of bacterial artificial chromosome (BAC) transgenic constructs is described. We give an overview of different transgenic inserts, such as fluorescent proteins (alone or in combination with Cre variants), diphtheria toxin receptor, lacZ, and light-activated ion channels. The most reliable and versatile approach to express these genes is by using BACs, which allow "highjacking" of the expression pattern of a gene without characterizing its transcriptional control elements. Here, we describe the necessary cloning techniques compared with conventional transgenesis. With the provided "toolbox" of already available transgene constructs, the generation of the BAC transgenes is made easy and rapid. We provide a comprehensive outline how to insert the different transgenes into a chosen BAC by either ET cloning or recombineering. We also describe in detail the methods to identify the correct insertion and the integrity of the final BAC construct, and finally, the preparation of the BAC DNA for oocyte injection is described. genesis 00:1-17, 2010. (c) 2010 Wiley-Liss, Inc.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:04 Faculty of Medicine > Institute of Pharmacology and Toxicology
04 Faculty of Medicine > Institute of Experimental Immunology
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Language:English
Date:April 2010
Deposited On:06 May 2010 11:08
Last Modified:05 Apr 2016 14:05
Publisher:Wiley-Blackwell
ISSN:1526-954X
Publisher DOI:10.1002/dvg.20612
PubMed ID:20143345
Permanent URL: http://doi.org/10.5167/uzh-33423

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