Js. Sandhu et al., Enhanced expression of the human respiratory syncytial virus-F gene in apple leaf protoplasts, PL CELL REP, 18(5), 1999, pp. 394-397
Expression of the human respiratory syncytial virus (RSV) fusion protein (F
) gene under the control of the cauliflower mosaic virus (CaMV) 35S promote
r was analyzed by enzyme-linked immunosorbent assay ( ELISA) in polyethylen
e glycol-transfected apple leaf protoplasts. In particular, we examined whe
ther RSV-F gene expression could be enhanced by addition of a viral leader
and a plant enhancer to the chimeric gene construct. Insertion of the 5'-un
translated leader from alfalfa mosaic virus (AMV) RNA 4 between the CaMV 35
S promoter and the RSV-F gene increased viral expression by 5.5-fold compar
ed to the construct without the leader. The addition of a transcriptional e
nhancer from the pea plastocyanin gene (PetE) upstream of the CaMV 35S prom
oter to a construct containing the AMV leader further increased RSV-F gene
expression by 1.4-fold. Immunoblot assays showed that the RSV-F expressed i
n transfected apple protoplasts reacted with RSV-F monoclonal antibodies an
d was of the expected molecular mass of 68 kDa. These results demonstrated
that the RSV-F recombinant protein was expressed in an antigenic form in pl
ant cells. Furthermore. protein expression was enhanced by modifying the tr
ansfection vector using both a leader and an enhancer linked to a promoter.