Inducible transgene expression systems represent a key methodology for studying gene expression and function. Direct, inducible control over transgene expression allows for developmental and time point studies in vivo. Developing mouse models with inducible expression systems, however, can be quite complicated and costly, and require mating with transcriptional activator or silencer mouse lines that might not already exist, when tissue-specificity is desired.
ingenious has taken the complicated process of generating tissue-specific inducible transgene expression systems and simplified it. By designing a targeting vector that contains a floxed stop cassette and all of the necessary Tet-On control components, we enable tissue-specificity and inducibility without the need for transactivator or silencer mouse lines. Simply mate the targeted transgenic line with a tissue-specific Cre recombinase mouse line of choice and introduce doxycycline for inducible/reversible expression of your transgene in your tissue of interest.
The design works by incorporating a reverse tetracycline-controlled transactivator (rtTA) component, a TRE3G (tetO) promoter, and a floxed Neomycin-Stop cassette into the “safe harbor” Rosa26 locus. The Neomycin-Stop cassette, inserted downstream of the CAG promoter, prevents rtTA expression, which is one of the key components required for transgene expression from the TRE3G promoter.
Mating the Inducible Rosa26-Express™ mouse line to a ubiquitous, or tissue-specific Cre recombinase mouse line allows for removal of the floxed Neomycin-Stop cassette, for expression of rtTA from the pCAG promoter.
Doxycycline introduced to the mouse via food, water, or injection, binds with high affinity to the rtTA, allowing it in turn to bind to the TRE3G promoter and activate expression of the transgene. Removal of doxycycline renders the rtTA unable to bind to the TRE3G promoter and expression stops.
1) Rapid-Rosa26™ Targeting technology from ingenious
2) Tet Systems: Principles and Components
3) Tet Systems: Home Page
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5) Stieger K, Belbellaa B, Le Guiner C, Moullier P, Rolling F. 2009. In vivo gene regulation using tetracycline-regulatable systems. Adv Drug Deliv Rev 61(7-8): 527-541.