Inducible Cre recombinase systems have been developed to bypass initial lethal phenotypes and to provide access to later embryonic or adult phenotypes. Here we described the generation of a transgenic recombinant mouse that combines a tetracycline dependent switch with generalized Cre recombinase expression by targeting the ubiquitously expressed ROSA26 locus. This transgenic strain (R26rtTA-TRECre) was developed using a universal and simplified gene delivery system designed to facilitate the generation of conditional animals by integrating both elements, the reverse tetracycline controlled trans-activator (rtTA) and rtTA inducible promoter in a single vector. In this transgenic strain, the endogenous ROSA26 promoter drives rtTA expression through a splice acceptor site. The tetracycline inducible promoter or tetracycline response element (TRE), cloned in opposite orientation to the ROSA26 locus and separated from the rtTA element by a 5kb human p53 intron, drives Cre recombinase expression. Crossing these mice with a Cre reporter strain, in which the reporter gene is activated by the elimination of a loxP flanked DNA sequence, showed that Cre DNA mediated recombination was ubiquitously and effectively induced during various prenatal developmental windows upon treatment with a tetracycline analog (doxycycline). Background Cre recombinase expression levels were observed in some tissues in the absence of the inducer, mostly during late embryonic developmental stages. Background recombination levels were low during development and most prominent in nervous tissue. Cre recombinase expression could not be effectively induced in adult animals. While rtTA mRNA levels were high in adult tissues, Cre recombinase mRNA levels remained low after doxycycline treatment. Therefore, the mouse strain described here provides a valuable tool to further analyze the function of genes during specific developmental windows, by allowing the effective inactivation of their function throughout defined stages of embryonic development. The mouse strain described here is currently being used in our lab to further analyze the function of genes during specific developmental windows, by allowing the effective inactivation of their function throughout defined stages of embryonic development. Specifically we are crossing this strain with sonic hedgehog (Shh) LoxP animals to further study the role of Shh on dopamine neuron development. As Shh KO animals die in utero, being able to delete Shh just before the appearance of TH in the ventral mesencephalon, may help us define if Shh is a mandatory factor for the development of DA neurons in vivo.
|Backman, Cristina M; Zhang, YaJun; Malik, Nasir et al. (2009) Generalized tetracycline induced Cre recombinase expression through the ROSA26 locus of recombinant mice. J Neurosci Methods 176:16-23|