Expression of Human Papillomavirus Type 16 L1 Protein in Transgenic Tobacco Plants

To develop a plant expression system for the production of the human papillomavirus type 16(HPV16)vaccine,we investigated whether the HPV16 L1 protein can be expressed in tobacco plants andwhether it can be used as the cheapest form of edible vaccine.The HPV16 L1 coding sequence was amplifiedby PCR using specific primers from the plasmid pGEM-T-HPV16 containing the template sequence,andsubcloned into the intermediate vector pUCmT and binary vector pBI121 consecutively to obtain the plantexpression plasmid pBI-L1.The T-DNA regions of the pBI-L1 binary vector contained the constitutiveCauliflower mosaic virus(CaMV)35S promoter and the neomycin phosphotransferase npt Ⅱ gene,whichallowed the selection of transformed plants using kanamycin.The tobacco plants were transformed by co-cultivating them,using the leaf disc method,with Agrobacterium tumefaciens LBA4404,which harbored theplant expression plasmid.The regenerated transgenic tobacco plants were selected using kanamycin,andconfirmed by PCR.The results of the Southern blot assay also showed that the HPV16 L1 gene was inte-grated stably into the genome of the transformed tobacco plants.The Western blot analysis showed that thetransformed tobacco leaves could express the HPV16 L1 protein.Furthermore,it was demonstrated byELISA assay that the expressed protein accounted for 0.034%-0.076% of the total soluble leaf protein,wasable to form 55 nm virus-like particles compatible with HPV virus-like particle(VLP),and induced mouseerythrocyte hemagglutination in vitro.The present results indicate that the HPV16 L1 protein can be ex-pressed in transgenic tobacco plants and the expressed protein possesses the natural features of the HPV16L1 protein,implying that the HPV16 L1 transgenic plants can be potentially used as an edible vaccine.