[Construction and identification of tetracycline-inducible rat Smad7 eukaryotic expression vector].

ABSTRACT

OBJECTIVE: To construct a tetracycline-inducible eukaryotic expression vector of rat Smad7. METHODS: The total RNA was extracted from normal rat kidney with Trizol agent. Rat Smad7 cDNA fragment was cloned by RT-PCR, and was inserted into the restriction site between Nhe I and Hind III of the inducible eukaryotic expression vector pBI-L by tetracycline. pBI-L-Smad7 was constructed by digestion and ligation, and detected by restriction endonuclease digestion and sequencing. RESULTS: The recombinant eukaryotic expression vector pBI-L-Smad7 was constructed correctly as confirmed by restriction endonuclease digestion and sequencing. The fragment of pBI-L-Smad7 digested with restriction endonucleases and the sequence of inserted Smad7 cDNA were consistent with the results of theoretical analysis. CONCLUSION: The tetracycline- inducible eukaryotic expression vector of rat Smad7, pBI-L-Smad7, is constructed successfully, which may facilitate further clinical study of Smad7 gene therapy for tissue and organ fibrosis.