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Oral immunization of animals with transgenic cherry tomatillo expressing HBsAg

机译：用表达HBsAg的转基因樱桃番茄对动物进行口服免疫

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AIM: To investigate the expression of recombinant HBsAg (rHBsAg) in transgenic cherry tomatilio in order to explore the feasibility of producing HBV oral vaccine with cherry tomatillo by animal immune tests. METHODS: The recombinant plant expression vector containing HBsAg gene was constructed. Mediated with Agrobacterium tumefaciens, HBsAg gene was transferred into cotyledons of cherry tomatillo. Transformed cherry tomatillos were obtained through hygromycin delay-selection. Integrated DNA in transgenic cherry tomatillo was confirmed by hygromycin resistance selection, Gus detection, polymerase chain reaction (PCR) and dot blotting analysis. Antigenicity of rHBsAg was examined by ELISA and the immunogenicity of rHBsAg derived from transgenic cherry tomatillo tissues was confirmed by oral feed of transformed tissues to BALB/c mice primed with commercial HBV vaccines. Specific antibody titers in mice's serum were examined by ELISA every week. RESULTS: By far, 10 positive lines of transgenic cherry tomatillos containing HBsAg gene were obtained. Among different organs of the same transgenic cherry tomatillo, level of rHBsAg expressed in leaves was the highest with the yield up to 300ng/g fresh weight. And the rHBsAg expression level in fruits was about 10 ng/g fresh weight. In animal immune tests, oral delivery with transgenic tissues tc mice primed with commercial vaccine instead of naive mice resulted in significant immune response. CONCLUSION: The result of this animal immune test indicated the rHBsAg derived from transgenic cherry tomatillo possessed normal immunogenicity. This work demonstrated the feasibility to generate oral immunogenic rHBsAg in transgenic cherry tomatillo, and would provide some experimental approach for the production of low-cost oral vaccines using transgenic cherry tomatillo in large scale.
机译：目的：研究重组HBsAg（rHBsAg）在转基因樱桃番茄中的表达，以探讨通过动物免疫试验生产樱桃番茄生产HBV口服疫苗的可行性。方法：构建含有HBsAg基因的重组植物表达载体。在根癌农杆菌的介导下，HBsAg基因被转移到樱桃番茄的子叶中。通过潮霉素延迟选择获得转化的樱桃番茄。通过潮霉素抗性选择，Gus检测，聚合酶链反应（PCR）和斑点印迹分析确认了转基因樱桃番茄中的整合DNA。通过ELISA检查了rHBsAg的抗原性，并且通过将转化的组织口服饲喂商业HBV疫苗接种的BALB / c小鼠，证实了来自转基因樱桃番茄组织的rHBsAg的免疫原性。每周通过ELISA检查小鼠血清中的特异性抗体滴度。结果：到目前为止，获得了含有HBsAg基因的10株转基因樱桃番茄阳性株。在同一转基因樱桃番茄的不同器官中，叶片中表达的rHBsAg水平最高，鲜重达300ng / g。水果中的rHBsAg表达水平约为10 ng / g鲜重。在动物免疫测试中，用商业疫苗而非天然小鼠接种的转基因组织tc小鼠口服给药可导致明显的免疫反应。结论：该动物免疫试验结果表明，转基因樱桃番茄的rHBsAg具有正常的免疫原性。这项工作证明了在转基因樱桃番茄中产生口服免疫原性rHBsAg的可行性，并将为大规模使用转基因樱桃番茄生产低成本口服疫苗提供一些实验方法。

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