Tet-On 3G Bidirectional Inducible Expression System

The Tet-On 3G systems allow inducible gene expression only in the presence of doxycycline (Dox)

The Tet-On 3G systems allow inducible gene expression only in the presence of doxycycline (Dox). When Dox binds, the transactivator undergoes a conformational change allowing it to bind tet operator (tetO) repeats within the TRE3G promoter.

Tet-On 3G inducible expression systems with CMV regulator vectors

Tet-On 3G inducible expression systems with CMV regulator vectors.

Doxycycline-regulated expression of two genes

Doxycycline-regulated expression of two genes. ZsGreen1 and firefly luciferase were cloned into separate multiple cloning sites that flank the inducible bidirectional promoter in the pTRE3G-BI vector. The plasmid was then cotransfected into HEK 293 cells with pCMV-Tet3G (which expresses the Tet-On 3G transactivator). Cells were treated with the indicated doses of doxycycline, and induced gene expression was measured 24 hr later. Induced expression of ZsGreen1 fluorescent protein was determined by fluorescent microscopy and luciferase activity was measured using a luminometer (RLU = Relative Light Units).

PTRE3G-BI inducible bidirectional promoter shows equivalent induced expression of two genes

PTRE3G-BI inducible bidirectional promoter shows equivalent induced expression of two genes. ZsGreen1 and mCherry fluorescent proteins were cloned into separate multiple cloning sites that flank the inducible bidirectional promoter in the pTRE3G-BI vector. The plasmid was then cotransfected into HEK 293 cells with pEF1a-Tet3G (which expresses the Tet-On 3G transactivator). Cells were treated with 100 ng/ml doxycycline and induced expression of the fluorescent proteins was observed 24 hr later. Cells in the mixed population that expressed high levels of ZsGreen1 also showed similarly high levels of mCherry expression, indicating equivalent expression from both sides of the bidirectional promoter.