Abstract
Expression of the human respiratory syncytial virus (RSV) fusion protein (F) gene under the control of the cauliflower mosaic virus (CaMV) 35S promoter was analyzed by enzyme-linked immunosorbent assay (ELISA) in polyethylene glycol-transfected apple leaf pro- toplasts. In particular, we examined whether RSV-F gene expression could be enhanced by addition of a viral leader and a plant enhancer to the chimeric gene construct. Inser- tion of the 5'-untranslated leader from alfalfa mosaic virus (AMV) RNA 4 between the CaMV 35S promoter and the RSV-F gene increased viral expression by 5.5-fold com- pared to the construct without the leader. The addition of a transcriptional enhancer from the pea plastocyanin gene (PetE) upstream of the CaMV 35S promoter to a con- struct containing the AMV leader further increased RSV-F gene expression by 1.4-fold. Immunoblot assays showed that the RSV-F expressed in transfected apple pro- toplasts reacted with RSV-F monoclonal antibodies and was of the expected molecular mass of 68 kDa. These re- sults demonstrated that the RSV-F recombinant protein was expressed in an antigenic form in plant cells. Further- more, protein expression was enhanced by modifying the transfection vector using both a leader and an enhancer linked to a promoter.
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