Construction and Identification of Hsa-microRNA-138 Lentiviral Vector

Objective: To construct hsa-microRNA-138 lentiviral vector.Methods: Pre-miR-138-2 amplified by PCR was cloned into plenti-GFP vector,and was identified by restriction endonuclease digestion and nucleotide sequencing,and then the lentivirus was packaged and virus titer was detected.After successfully constructed,the vector was transfected into human pancreatic cancer cell PANC-1.After 48h,the expression of hsa-miR-138 was detected by Real-time Q-PCR.Results: Restriction enzyme digestion and DNA sequencing demonstrated that the inserted sequences were correct.The titer of virus was 1×109 TU/mL.After lentiviral infection,green fluorescence was detectable under fluorescence inverted microscope in PANC-1 cells,and real-time Q-PCR showed that expression amount of hsa-miR-138 in transfected cells was more than that in non-transfected cells.Conclusions: The lentiviral vector expressing hsa-miR-138 efficiently and stably is successfully constructed.