Construction of shRNA lentiviral vector targeting human NOB1 gene and identification of RNAi efficiency

Objective: To construct the shRNA lentiviral vectors targeting human NOB1 gene and detect its effect of gene silence in SKOV3 and HEY cells. Methods: The specific siRNA sequences targeting human NOB1 gene were cloned into pLVTHM lentiviral vector. After screening for the valid siRNA, the lentivirus particles were packaged and NOBl specific shRNA was transmitted into SKOV3 and HEY cells. Then, real-time PCR was performed to determine the expression level of NOB1. Results: PCR and sequencing results revealed that shRNA plasmids were correctly constructed. The virus with a titer of 8 X 10 8 PU·L -1 was successfully packaged. The NOB1 expressions in SKOV3 and HEY cells were down-regulated at mRNA level by virus infection. The expression levels of NOB1 mRNA was decreased by 73.5% in SKOV3 and by 82.2% in HEY cells compared with negative control. Conclusion: The recombinant lentiviral shRNA expression vector targeting human NOB1 gene has been successfully constructed. NOB1 mRNA can be down-regulated availably in SKOV3 and HEY cells.