A Transposon-Mediated System for Flexible Control of Transgene Expression in Stem and Progenitor-Derived Lineages

• pB-Tet-GOI features the latest generation of Tet transactivators (tTAs) and variants
• piggyBac transposition allows for genomic insertion of pb-Tet-GOI
• pb-Tet-GOI provides flexible control of transgenes in vitro and in vivo
• tTA variants permit reversible, constitutive, or induced constitutive expression

SummaryPrecise methods for transgene regulation are important to study signaling pathways and cell lineages in biological systems where gene function is often recycled within and across lineages. We engineered a genetic toolset for flexible transgene regulation in these diverse cellular contexts. Specifically, we created an optimized piggyBac transposon-based system, allowing for the facile generation of stably transduced cell lineages in vivo and in vitro. The system, termed pB-Tet-GOI (piggyBac-transposable tetracycline transactivator-mediated flexible expression of a genetic element of interest), incorporates the latest generation of tetracycline (Tet) transactivator and reverse Tet transactivator variants—along with engineered mutants—in order to provide regulated transgene expression upon addition or removal of doxycycline (dox). Altogether, the flexibility of the system allows for dox-induced, dox-suppressed, dox-resistant (i.e., constitutive), and dox-induced/constitutive regulation of transgenes. This versatile strategy provides reversible temporal regulation of transgenes with robust inducibility and minimal leakiness.

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