Construction of lentivirus-mediated multi-shRNAs vector and evaluation of anti-FMDV in vitro and vivo

【Objective】 The study was conducted to investigate the inhibit replication of foot-and-mouth disease virus (FMDV) by multi-shRNAs expression.【Method】 Three shRNAs were designed and chemically synthesized according to the conservative area in 3B and 3D regions of FMDV; induced by three different promoters respectively, they were constructed into a multi-shRNAs expressing lentiviral plasmid. The anti-FMDV multi-shRNAs expressing lentiviral particles were packaged by co-transfecting the three plasmid lentivirus packaging system into 293T cells. Infected FMDV into lentivirus-treated cells and suckling mice, inhibitions of FMDV were observed.【Result and conclusion】 Results showed that transgenic BHK-21 cells were obtained by infecting lentivirus. The expression of shRNAs in transgenic cells was detected by stem-loop RT-PCR. Inoculated with FMDV type O, the transgenic cells were proven to have an obvious inhibition to FMDV replication, which could reduce virus growth by three folds (24 h post-infection). After infection of FMDV type O strain into 3-5 days suckling mice, no mouse mortality was observed under 5 LD₅₀ titer, and survival time of the dead mice extended compared with negative control under 20 LD₅₀ titer. Based on the above results, it can be concluded that the anti-FMDV multi-shRNAs expressing lentiviral vector can improve FMDV resistance of BHK-21 cells and suckling mice.