Cloning Of Hepatitis B Surface Antigen S Gene And Tissue Culture Of Cucumber For Gene Transformation

Hepatitis B is one of the most dangerous infectious diseases that do harm to people's health and it ravages in almost everywhere on the earth. Widely vaccination for hepatitis B has been the only economical and effective means to control the disease. And it is one of the most crucial tasks for us in the near future to find out how to make vaccine more safe, economical, effective and easy to application.With the rapid development of molecular biology and gene engineering, plants can be used as the bioreactor for proteins, carbohydrates and lipids. Subtype adr is the prevailing subtype of HBV in Chinese people. We used the coding sequence of this subtype searched from genebank as the model of PCR reaction and designed a pair of primer: 5'-cgggatccatggagaacacaacatcagga-3' (Forward primer), 5'-cgagctcttaaatgtatac ccaaagaca-3' (Backward primer). The DNA prepared from the serum of HBV sufferers was amplified by PCR and the S gene of HBsAg was then obtained. After purified and digested by restricted enzymes, the S gene of HBsAg was successfully cloned into pBI121, which was a kind of plant expression vector. The recombinant plasmid was transformed into E. coli strain JM109 and then transformed into Agrobacterium tumefaciens LBA4404 by heat shock, which can be used for plant transformation. In the same time, we had an attempt in tissue culture of cucumber (Cucumis sativus L.) in two different ways and the effect of the explant tissue type, age of explant and hormone ratio in the medium on callus induction was investigated. We found that buds directly regenerated from cotyledon explants is suitable for plant transformation due to its high regenerating rate and quickly seedling, which established a basis for further study on the transformation of cucumber.S gene of HBV subtype adr used in this research is the prevailing subtype in Chinese people, which make it more aiming in the precaution of HBV. This work can have certain impact on the development of cucumber-derived HBV vaccine and make a change in immunological concept and an improvement in the traditional immunological methods.