Molecular Farming, an Effective System for the Production of Immunogenic Crimean-Congo Hemorrhagic Fever Virus Glycoprotein

The main aim of this study was to obtain the Crimean-Congo hemorrhagic fever virus (CCHFV) glycoprotein, through either stable transgenic plants or using a transient expression system, and determine the yield, quality and finally the immunogenicity of the plant-made CCHFV glycoprotein in a mouse model. We designed and synthesized a codon-optimized G1/G2 gene from the G1 and G2 parts of the CCHFV glycoprotein by bioinformatic analysis. The synthetic construct was cloned into a plant expression vector and tobacco plants were both transiently and stably transformed. The transgenic plantlets or tobacco-derived hairy roots confirmed by PCR and Southern blot analyses. The intact 98 kDa G1/G2 glycoprotein was produced by a transient expression system at as much as 3.3 mg/kg fresh weight. The recombinant G1/G2 protein was analyzed in stable lines by G1/G2 ELISA and Western blot. The yield in the transgenic hairy root line was significantly higher than that in transgenic tobacco lines. Finally, the immunogenicity of the plant-made G1/G2 glycoprotein was evaluated by its subcutaneous administration in mice when compared with positive and negative controls. This plant-purified G1/G2 protein produced a high titer of anti-CCHFV glycoprotein IgG antibodies in mice.