Objective    For the research of the molecular mechanism of TCR process

we constructed a bidirectional Tet-responsive promoter plasmid including two report gene luciferase and SupF gene. This plasmid was used to detect the mutation frequency and spectra mediated by cisplatin in the TCR pathway.      Methods    SupF gene was cloned into a Tet-responsive plamid PBI-L that contains a bidirectional Tet-responsive promoter

and this subclone was named pTCR-1. Then SV40 ori fragment obtained form pEGFP-1 plasmid was ligated with pTCR-1

and named pTCR-2. The pTCR-2 plasmid was exposed to cisplatin in vitro and transfected into Tet-on 293 cell lines

then cultured in the media containing DOX for 48 h. The mutation frequency and spectra of supF gene were finally detected.     Results    The restriction digestion and sequence analysis demonstrated that the length of pTCR-2 plasmid and the SupF gene sequence were correct. The mutation frequency and spectra were obtained at the present of DOX.     Conclusion    A new Tet-responsive promoter plasmid has been successfully established and works as expected. It can be used to analyze the mutation frequency and spectra induced by cisplatin.