Expression And Functional Analysis Of BjMT2, A Metallothionein Type2 From Brassica Juncea, In Tobacco Plant

For characteristic and functional studies of BjMT-2, a plant metallothionein type2 from Brassica juncea, this research work was focused on the following four aspacts: 1) Transformation of plant metallothionein type2 cDNA clone gene into tobacco leaves via Agrobacterium tumefacience-mediated transformation. 2) The factors affected the plant regeneration and the transformation of BjMT-2 into tobacco leaves includied the hormone application, antibiotic concentration and Agrobacterium mediated transformation were analized, which helped to establish a new modified system for BjMT-2 gene transformation into tobacco leaves. 3) Confirmation of BjMT-2 gene transformation and expression by PCR and Northern blot analyses. 4) Immunodetection of BjMT-2, which was first applied in this research field, by Western blot analysis using the antibody prepared by a tandemly arranged BjMT-2 (BjMT-2:: BjMT-2) gene expressed in E. coli directed against the full lenth of amino acid in protein. This antibody would be an useful tool for detection of MT-2 in or among different plant species or specific tissue, respectively.In the present research it is confirmed: The level of GSH and chlorolophyll in transformed and wild type of tobacco leaves did not show significant variation but increased in the transformed leaves after 100 mM Cu²⁺ stress; The ATP hydrolysis showed that the PATPase activities were higher in the microsomal fraction prepared from transformed tobacco leaves than those from wild types. However, the V-ATPase activities were declined in comparison with the wild types; The transgenic plants differentiated in their phenotypes during their initial growing and in root elongation compared with those of wild types. Those above results were lack or neglected in the previous researchs.