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. 2017 Feb 12;7(4):912–925. doi: 10.7150/thno.16615

Figure 2.

Figure 2

Genetic stability of Rluc-JEV in BHK-21 cells. (A) The Rluc-JEV virus (P0) was blindly passaged on BHK-21 cells for five rounds (P1-P5). Luciferase activity of each passaged viruses was detected as described in Materials and methods. Error bars represent the standard deviation of triplicate measurement. *, P<0.05; ***, P<0.001; ****, P<0.0001. (B) Detection of the Rluc gene during blind passage. Viral RNA was extracted from the supernatants of BHK-21 cells infected with each passaged virus, and RT-PCR assay was performed to amplify the region from 5'UTR to prM which includes the Rluc gene. (C) Individual Rluc-JEV viruses were isolated by six rounds of plaque purification, designated as P6 viruses. P6 viruses were then serially passaged on BHK-21 cells for another three rounds (P7-P9). Luciferase activity in the cells infected with P7-P9 viruses was detected. Error bars represent the standard deviation of triplicate measurement. (D) Detection of the Rluc gene of P7-P9 viruses by RT-PCR using the same primers in (B). The expected band, corresponding to 1.6 kb, indicates the presence of the Rluc gene in the passaged plaque-purified reporter viruses.