Production and Characterization of Hepatitis B Recombinant Vaccine in Tobacco (Nicotiana tabacum cv. ‘Kanchun’)

Immunization with Hepatitis B vaccine is the most effective means of preventing Hepatitis B virus infection and its consequences. Plants are a potential source of Hepatitis B surface Antigen (HBsAg) that is not dependent upon process technology to ensure protein folding and particle assembly. A plant-based HBsAg expression system makes possible the testing of an oral immunization strategy by simply feeding plant samples. The primary means of transformation is Agrobacterium-mediated gene transfer which has provided a reliable means of creating transformants in a wide variety of species and also can express important pharmaceutical products. Leaf explants of tobacco were transformed with the Hepatitis B surface antigen gene along with nptII as an antibiotic selection marker gene. The presence of the HBsAg gene in putative transformants was confirmed by the presence of a 900-bp band in PCR analysis. The crude protein obtained from the transformed tobacco plants was tested by SDS-PAGE for the presence of a 24 kDa protein. Western-blot and ELISA confirmed the antigen specificity and immunogenic nature of the Hepatitis B surface antigen S-protein. T1 generation seeds obtained from transgenic tobacco plants were tested for inheritance analysis by germination in the presence of 100 ppm kanamycin. These showed a 3: 1 segregation ratio indicating Mendelian inheritance. Transgenic plants hold promise as low-cost vaccine production systems and this study emphasizes the integration and stability of recombinant protein expressed in tobacco plants.