Fig. 4.
Generation and characterization of complemented-defective OHFV-ΔNS1-Gluc virus. A Schematic of construction of OHFV-ΔNS1-Gluc clone. The Gluc2A sequences were inserted to replace the NS1 gene. B Generation of complemented OHFV-ΔNS1-Gluc virus. OHFV-ΔNS1-Gluc RNA was transfected into BHK21NS1 cells and NS3 expression was analyzed by IFA at different times post-transfection. C Comparison of growth kinetics between complemented OHFV-ΔNS1 and OHFV-ΔNS1-Gluc virus in BHK21NS1 cells. BHK21NS1 cells were infected at MOI of 0.05 IU/cell. Supernatants at indicated times were harvested and titers were determined as described in “Materials and Methods” section. D Correlation between viral titer and Gluc signals of defective OHFV-ΔNS1-Gluc virus in BHK21NS1 cells. BHK21NS1 cells were infected with OHFV-ΔNS1-Gluc virus at MOI of 0.05 IU/cell in a 12-well format. Supernatants and cell lysates were collected at indicated times. The titer of supernatants and Gluc signals of cell lysates were determined as described in “Materials and Methods” section. The data shown are the mean values of triplicate measurements with error bars showing standard deviation. E Correlation between Gluc activity and viral load of OHFV-ΔNS1-Gluc virus in BHK21NS1 cells. BHK21NS1 cells were infected with OHFV-ΔNS1-Gluc virus at MOI of 0.01, 0.02, 0.05, 0.1, 0.2, and 0.5 IU/cell in a 12-well format. At 20 hpi, cell lysates were collected and analyzed for Gluc activity. The data shown are the mean values of triplicate measurements with error bars showing standard deviation.