Expression of a consensus dengue virus envelope protein domain III in transgenic callus of rice

An effective dengue vaccine should elicit immune responses against all four different dengue virus serotypes. This study optimized the codon usage of a gene encoding consensus dengue virus envelope protein domain III (cEDIII) with cross-neutralizing activity against four dengue virus serotypes for plant expression. Then, a plant expression vector was constructed with this gene under the control of the rice amylase 3D promoter (RAmy3D), which is a strong inducible promoter under sugar starvation conditions. The synthetic cEDIII gene was fused with the RAmy3D signal peptide and ER retention signal, SEKDEL, and was introduced into rice callus by particle bombardment-mediated transformation. The integration and expression of cEDIII gene in transgenic rice callus was confirmed by genomic DNA PCR amplification, Northern blot analysis, and western blot analysis, respectively. Densitometric analysis determined that the highest expression level of the cEDIII protein in lyophilized rice callus was approximately 0.45 mg g⁻¹. These results suggest that it is feasible to use transgenic rice callus to produce the consensus dengue virus envelop protein domain III for edible vaccine purposes.