FIG 2.

Confirmation of capsid deletion in infectious ΔC-CHIKV mutants. (A and B) Detection of viral protein expression (nsP2, E1, E2, and capsid) in infected cells by IFAs and Western blot assays. BHK-21 cells were infected with the supernatant that was collected from either WT CHIKV or ΔC-CHIKV RNA-transfected BHK-21 cells at 72 hpt. At 72 hpi, cells were subjected to IFAs (A) and Western blot assays (B) to analyze the expression of different viral proteins. (C) Detection of the expression of the capsid gene in infected cells and culture media by an RT-PCR assay. Viral RNAs were extracted from infected cells and culture media. RT-PCR was performed with the primer pair spanning the nsP4-E3 region. The resulting RT-PCR products were resolved by 1% agarose gel electrophoresis. The expected 1.7-kb and 1.0-kb bands were observed for WT CHIKV and ΔC-CHIKV, respectively. (D) Sequence chromatograms of the N terminus of capsid. (E) Effects of different agents on the infectivity of infectious WT CHIKV or ΔC-CHIKV. WT CHIKV-eGFP or ΔC-CHIKV-eGFP (MOI = 0.01) was incubated with 5 μg/ml RNase A during 20 min at 25°C or with 0.1% Triton X-100 during 1 h at 25°C, respectively. The percent infectivity in each case was calculated by dividing the infectivity of untreated samples (No treat). The data are the means ± standard deviations (SD) from at least three independent experiments.