title: Expression of avian reovirus minor capsid protein in plants.
abstract: The minor coat protein of the avian reovirus (ARV), σC, encoded by the S1 genome segment, is one of the major candidates for the development of a subunit vaccine against ARV infection. To develop a plant-based vaccine to immunize poultry against ARV infection, we constructed 4 plant nuclear expression vectors with or without codon modification of the S1 gene, and their expression was driven by a CaMV 35S promoter or rice actin1 promoter. In addition, the expressed σC proteins were targeted subcellularly to cytosol or chloroplasts, respectively. Agrobacterium containing the S1 expression constructs was used to transform tobacco leaf disks, and transformants were selected with kanamycin (100 μg/ml). The integration of the S1 transgene into the tobacco chromosome was confirmed by PCR and Southern blot analysis. Western blot analysis with antiserum against σC was performed to determine the expression of σC protein in transgenic tobacco plants. The highest expression levels of σC protein in the cellular extracts of selected p35S1, pActS1 and p35UmS1 transgenic lines were 0.013%, 0.021% and 0.0013% of the total soluble protein, respectively, but the protein was barely detectable in p35TmS1 transgenic lines. However, the level of σC protein expression was not associated with the level of corresponding RNA transcripts in selected transgenic lines. Taken together, the results suggest that the major limiting factor for the expression of σC protein in plants might be at the post-transcriptional level.
authors.affiliation: Institute of Biotechnology, National Cheng Kung University, Tainan 701, Taiwan.
authors.forename: Shih-Wei
authors.initials: SW
authors.lastname: Lu
authors.forename: Kuo-Chih
authors.initials: KC
authors.lastname: Wang
authors.forename: Hung-Jen
authors.initials: HJ
authors.lastname: Liu
authors.forename: Ching-Dong
authors.initials: CD
authors.lastname: Chang
authors.forename: Hao-Jen
authors.initials: HJ
authors.lastname: Huang
authors.forename: Ching-Chun
authors.initials: CC
authors.lastname: Chang
mesh_terms: D000956:Antigens, Viral / Q000096:biosynthesis* / Q000235:genetics; D015139:Blotting, Southern; D015153:Blotting, Western; D036022:Capsid Proteins / Q000096:biosynthesis* / Q000235:genetics; D005822:Genetic Vectors; D016254:Mutagenesis, Insertional; D030101:Orthoreovirus, Avian / Q000235:genetics*; D030821:Plants, Genetically Modified / Q000235:genetics / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D011995:Recombination, Genetic; D012231:Rhizobium / Q000235:genetics; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism; D014170:Transformation, Genetic; D014765:Viral Vaccines / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D000956:Antigens, Viral; D036022:Capsid Proteins; D011994:Recombinant Proteins; D014765:Viral Vaccines