title: Contained and high-level production of recombinant protein in plant chloroplasts using a temporary immersion bioreactor.
abstract: Chloroplast transformation is a promising approach for the commercial production of recombinant proteins in plants. However, gene containment still remains an issue for the large-scale cultivation of transplastomic plants in the field. Here, we have evaluated the potential of using tobacco transplastomic cell suspensions for the fully contained production of a modified form of the green fluorescent protein (GFP+) and, a vaccine antigen, fragment C of tetanus toxin (TetC). Expression of these proteins in cell suspension cultures (and calli) was much less than in leaves, reaching 0.5%-1.5% of total soluble protein (TSP), but still produced 2.4-7.2 mg/L of liquid culture. Much better expression levels were achieved with a novel protein production platform in which transgenic cell suspension cultures were placed in a temporary immersion bioreactor in the presence of Thidiazuron to initiate shoot formation. GFP+ yield reached 660 mg/L of bioreactor (33% TSP), and TetC accumulated to about 95 mg/L (8% TSP). This new production platform, combining the rapid generation of transplastomic cell suspension cultures and the use of temporary immersion bioreactors, is a promising route for the fully contained low-cost production of recombinant proteins in chloroplasts.
authors.affiliation: Division of Biology, Faculty of Natural Sciences, Imperial College London, London, UK. franck.michoux@imperial.ac.uk
authors.forename: Franck
authors.initials: F
authors.lastname: Michoux
authors.forename: Niaz
authors.initials: N
authors.lastname: Ahmad
authors.forename: James
authors.initials: J
authors.lastname: McCarthy
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Nixon
mesh_terms: D001702:Biopharmaceutics / Q000379:methods; D019149:Bioreactors; D002736:Chloroplasts / Q000235:genetics / Q000378:metabolism*; D018506:Gene Expression Regulation, Plant; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics; D018515:Plant Leaves / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics; D013744:Tetanus Toxin / Q000096:biosynthesis / Q000235:genetics; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D013744:Tetanus Toxin; D049452:Green Fluorescent Proteins