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

Torbjörn Johansson, Ilja Broll, Theresa Frenz, Saskia Hemmers, Burkhard Becher, Hanns Ulrich Zeilhofer, Thorsten Buch

Research output: Contribution to journalArticlepeer-review

29 Scopus citations

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 trans-gene 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 48: 264-280, 2010.

Original languageEnglish
Pages (from-to)264-280
Number of pages17
JournalGenesis
Volume48
Issue number4
DOIs
StatePublished - Apr 2010
Externally publishedYes

Keywords

  • Bacterial artificial chromosome
  • Channelrhodopsin
  • Cre
  • CreER
  • EGFP
  • ET recombination
  • Keywords transgenic mouse
  • LacZ
  • Recombineering
  • TdTomato

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