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. 2002 Feb;76(3):1422–1434. doi: 10.1128/JVI.76.3.1422-1434.2002

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Copyright © 2002, American Society for Microbiology

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FIG. 4. — Strategy to assemble TGEV-Rep(AvrII) VRPs. (a) In the full-length TGEV-Rep(AvrII) cDNA construct, ORF 3A has been replaced with GFP, and ORF 3B and the 5′ end of the E gene have been deleted. To produce packaged replicon particles, replicon RNA-transfected cells were infected with VEE VRPs expressing the TGEV E protein [VEE-TGEV(E)]. Alternatively, TGEV-Rep(AvrII) replicon RNAs can be coelectroporated with pVR21-E1-derived transcripts. TGEV VRPs should be released from cells that can be used as single-hit expression vectors. (b) Cultures of ST cells were infected with either wild-type (wt) TGEV alone or with TGEV and VEE VRPs expressing a G₁ Norwalk-like virus capsid (wt + VEE) at an MOI of 5 for 1 h at room temperature. The inocula were removed, and the cultures were incubated in complete medium at 37°C. Samples were harvested at the indicated times and assayed by plaque assay in ST cells.

FIG. 4. — Strategy to assemble TGEV-Rep(AvrII) VRPs. (a) In the full-length TGEV-Rep(AvrII) cDNA construct, ORF 3A has been replaced with GFP, and ORF 3B and the 5′ end of the E gene have been deleted. To produce packaged replicon particles, replicon RNA-transfected cells were infected with VEE VRPs expressing the TGEV E protein [VEE-TGEV(E)]. Alternatively, TGEV-Rep(AvrII) replicon RNAs can be coelectroporated with pVR21-E1-derived transcripts. TGEV VRPs should be released from cells that can be used as single-hit expression vectors. (b) Cultures of ST cells were infected with either wild-type (wt) TGEV alone or with TGEV and VEE VRPs expressing a G₁ Norwalk-like virus capsid (wt + VEE) at an MOI of 5 for 1 h at room temperature. The inocula were removed, and the cultures were incubated in complete medium at 37°C. Samples were harvested at the indicated times and assayed by plaque assay in ST cells.