Enhanced expression of the human respiratory syncytial virus-F gene in apple leaf protoplasts

J. S. Sandhu, M. D. Osadjan, S. F. Krasnyanski, L. L. Domier, S. S. Korban, D. E. Buetow

Research output: Contribution to journalArticlepeer-review

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 protoplasts. 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. Insertion 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 compared 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 construct 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 protoplasts reacted with RSV-F monoclonal antibodies and 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 plant cells. Furthermore, protein expression was enhanced by modifying the transfection vector using both a leader and an enhancer linked to a promoter.

Original languageEnglish (US)
Pages (from-to)394-397
Number of pages4
JournalPlant Cell Reports
Volume18
Issue number5
DOIs
StatePublished - Jan 1 1999

Keywords

  • Apple protoplasts
  • ELISA
  • Immunoblotting
  • RSV-F expression

ASJC Scopus subject areas

  • Agronomy and Crop Science
  • Plant Science

Fingerprint Dive into the research topics of 'Enhanced expression of the human respiratory syncytial virus-F gene in apple leaf protoplasts'. Together they form a unique fingerprint.

Cite this