Abstract
The dengue virus envelope glycoprotein (DV-E) has been identified as a promising candidate for the development of a subunit vaccine and to provide an antigen for diagnostic kits. In this study, cell suspension cultures of Nicotiana tabacum and Morinda citrifolia were transformed and evaluated for production of the DV-E serotype 2 protein. The expression cassette consisted of the DV-E gene along with a signal peptide at its 5′ end. In addition, a KDEL endoplasmic retention sequence was included in a second construct to evaluate its influence on accumulation levels of the recombinant protein. Expression cassettes were sub-cloned into a binary vector pCAMBIA 1305.2, and transformation was carried out using Agrobacterium tumefaciens. Transformed cell lines of both N. tabacum and M. citrifolia accumulated DV-E protein, although those carrying the construct containing the KDEL sequence showed higher accumulation levels than those without. Overall, cell suspension cultures of N. tabacum were more suitable for dengue antigen production than those of M. citrifolia. The highest levels of protein accumulation (0.71 ± 0.06 mg DV-E/L) were observed in tobacco cell lines at 5 days of culture, corresponding to 0.3% of total soluble protein. The recombinant protein was reactive with anti-E antibodies.
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Acknowledgments
CAM is supported by Universidad de Buenos Aires. AMG and JRT are supported by CONICET (Consejo Nacional de Investigaciones Científicas y Tecnológicas). CAM, AMG and JRT thank Dr. Andrea Gamarnik for giving us the DNA genetic sequences and her assistance and to CAMBIA research organization for supplying pCAMBIA vector. This work was granted by PICT 14-15112 and 14-33166, Agencia Nacional de Promoción Científica y Tecnológica, Argentina (ANPCyT), and UBACyT B111, Universidad de Buenos Aires.
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Martínez, C.A., Giulietti, A.M. & Rodríguez Talou, J. Expression of a KDEL-tagged dengue virus protein in cell suspension cultures of Nicotiana tabacum and Morinda citrifolia . Plant Cell Tiss Organ Cult 107, 91–100 (2011). https://doi.org/10.1007/s11240-011-9960-4
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DOI: https://doi.org/10.1007/s11240-011-9960-4