The Transformation And Expression Of HBsAg Gene In Tomato

In this experiment, a new plant expression vector, pCAMBIA1301/HB, was constructed, which was recombined with an exterior target gene of HBsAg. And, the HBsAg gene was transformed into tomato explant by agrobacterium LBA4404. Further, clump buds were induced from explants and took roots in given culture medium. The following particular results appeared in this experiment:1. Transformed agrobacterium LBA4404 with vector pCAMBIA1301/HB and spreaded them on a culture medium containing Kanamycin (Km) and Rifamycin (Rif). The grown bacteriums were confirmed by PCR assay and double enzymes digested assay that they were recombinants. The recombinants were given a name of LBA4404/pCAMBIA/HB.2. Clump buds were induced when tender tomato explants were put on the culture medium of MS+6-BA_2.0+IAA_0.2 and took roots normally on culture medium of 1/2MS+Hyg₂₅. At the same time , experiment also showed that no any clump bud could be induced again while medium contained a concentration of 25mg/L Hyg, and the roots could be restrained by a even lower concentration of 5mg/L Hyg, either.3. Transformed plants could take roots well in a high concentration of 25mg/L Hyg. Accommodating for two weeks, they were transplanted successfully to extraventricular flowerpots, and had a livability of 100%.4. Tomato explants were dipped with LBA4404/pCAMBIA/HB and put on filtrating culture medium to filtrate transgenic plants . About 6⁷ weeks later,new regenerations appeared. PCR and southern-blot assays showed that the regenerations were all transgenic tomato plants.5. Extracted protein of transgenic tomato plants and did western-blot & ELISA assays. Experiment results showed that the HBsAg gene had expressed in those tomato plants exactly, and the protein provided with immunocompetence.