pTRE2hyg is a response plasmid that expresses a gene of interest (Gene X) in CLONTECH's Tet-On™ and Tet-Off™ Gene Expression Systems and Tet-On and Tet-Off Cell Lines (1). The Tet Expression Systems and Cell Lines give researchers ready access to the tetracycline-regulated expression systems described by Gossen & Bujard (2; Tet-Off) and Gossen et al. (3; Tet-On). pTRE2hyg contains an MCS immediately downstream of the Tet-responsive PhCMV*-1 promoter. cDNAs or genes inserted into the MCS will be responsive to the tTA and rtTA regulatory proteins in the Tet-Off and Tet-On systems, respectively. PhCMV*-1 contains the Tet response element (TRE), which consists of seven copies of the 19-bp tet operator sequence (tetO). The TRE element is just upstream of the minimal CMV promoter (Pmin CMV), which lacks the enhancer that is part of the complete CMV promoter. Consequently, PhCMV*-1 is silent in the absence of binding of TetR or rTetR to the tetO sequences. Note that the cloned insert must have an initiating ATG codon. In some cases, addition of a Kozak consensus ribosome binding site (4) may improve expression levels; however, many cDNAs have been efficiently expressed in Tet systems without the addition of a Kozak sequence. pTRE2hyg also contains the hygromycin resistance gene for direct selection of stable transformants. The parental vector pTRE2 was originally described as pUHD10-3 in reference 5.
The pTRE2hyg-Luc Control Vector, packaged with the pTRE2hyg Vector, contains an additional 1649 bp encoding firefly luciferase inserted into the MCS. This vector can be used as a reporter of induction efficiency using standard luciferase detection reagents. It is not intended as a cloning vector.