Study Of Joint Intervention Of E-cadherin And Bmi-1 Genes Mediated By TALEN In Nasopharyngeal Carcinoma Metastasis In Vitro

Purposes: The aim of this study was to establish TALEN-mediated E-cad and Bmi-1 intervention of the metastasis of nasopharyngeal carcinoma cell line CNE-2,providing experimental basis for gene therapy of nasopharyngeal carcinoma.Methods: On the basis of the common multi-locus gene targeting vector pUC-DS1-DS2 constructed by our laboratory,the E-cad and Bmi-1 shRN A expression element were inserted by In-fusion technique to construct multi-locus gene targeting vectors pUC-DS1-CMV-E-cad-2A-Neo-DS2,pUC-DS1-Bmi-1 shRNA-Zeo-DS2.The E-cad and Bmi-1 shRNA gene targeting vectors were transferred with TALEN to CNE-2 cells individually or simultaneously.The integration of target genes were detected by PCR,the expression of E-cad,Bmi-1 and marker protein(Vimentin,?-1-Catenin,Fibronectin)of epithelial-mesenchymal transition(EMT)were detected by Western Blot.The effects of E-cad and Bmi-1 intervention on CNE-2 were measured by Transwell chamber,CCK-8 and flow cytometry.Results: The E-cad and Bmi-1 shRNA expression elements were successfully integrated into the genome of CNE-2 cells;the protein expression levels of E-cad was up-regulated,the protein expression levels of Bmi-1 was down-regulated.Compared with the control group,the migration and invasion ability of single gene or double gene transfection group were decreased significantly,the expression maker protein Vimentin,Fibronectin and ?-1-Catenin were down-regulated and up-regulated respectively of EMT.Furthermore,double gene intervention could inhibit the migration ability and EMT process of nasopharyngeal carcinoma cells significantly compared with single gene intervention.Conclusions: The study combined TALEN technique with gene targeting technique to intervent the metastasis of nasopharyngeal carcinoma in vitro,which will establish a key technique for gene therapy and animal genome editing.