摘要
AIM: To investigate the expression of recombinant HBsAg (rHBsAg) in transgenic cherry tomatillo 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 to 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.
AIM:To investigate the expression of recombinant HBsAg (rHBsAg) in transgenic cherry tomatillo 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 to 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.
基金
the grant from Natural Scientific Fundation of Fujian Province,No.C9910004
Xiamen Kaili Biologic Product Limited Company
