A Primary Study On Construction Of Plant Expression Vectors For The MT Gene Of Mice And Transformation To Aquatic Canna

Waste water containing heavy metals is discharged far more than ever before with industrialization, and it is a serious threat for the growth of crops and the human health. Heavy metals is difficult to discharged and controlled. Consequently,it is attentted more and more on ecological, environment, medicine, agriculture that how to discharge heavy metal pollution and its hazards .The author constructed the plant expression vector with MT gene which Combined with heavy metals only in the mammal. And choiced the plant transfer which grew in the poor environment or on the pollution of wetlands, The author tried to transfer MT (αα) gene into the plant by genetic engineering technology for the gene plant. The study has very important implications for improving the regional environment,the protection of ecological environment and the protection of animal and plant health .Construction of Plant Expression Vectors: In order to removed the gene because of the potential herbicide biological pollution problems and a more reasonable choice of the selection gene and reporter gene. The MT gene cassettes was cloned to the plant expression vector pCAMBIAl30l by gene technology Which were included in the expression of 35S-35S promoter, AMV, Tnos, It was used HPT gene replaced PPT gene and increased GUS reporter gene. At last, the MT gene plant expression vector pCAMBIAl301-MT was constructed, and then Transformed into LBA4404 for importing aquatic Canna genome.Preparation of aseptic seedling: the wild aquatic Canna was used as the materials of this experiment. They were rinsed with flowing water about 20 minutes after ablution, then cut the buds 1-2 cm long, immersed into 75% alcohol about 30 seconds under the sterile environment, put into 0.1%HgCl2 about 10 minutes, In the end, cut about 5mm3 stem apex with 3-4 leaf primordium as explants, and inoculated in proper nutrient medium after emundationing 4-6 times with SW. Tissue culture: aquatic Canna was used as experimental material. In the study of the induction conditions of multiple buds and callus, different explants of aquatic Canna were inoculated into different media which were added into different types and concentration of exogenous phytohormone. The results showed that: the optimal medium of Canna adventitious tip was MS+6-BA3mg/L + NAA0.1mg/L.; the optimal medium of Canna adventitious root was MS+NAA0.5mg/L; the optimal medium of Canna adventitious bud proliferation was MS+6-BA5mg/L + NAA0.2mg/L.Optimization of the system of genetic transformation: Taking aquatic Canna growth point as materials, we studied several factors of influencing Agrobacterium- mediated transformation system of aquatic Canna growth point, and drew the Optimal conditions: bacterial concentration OD600=0.3-05, AS 100μmol/L, infection time 15 min, and co-culture time 2 days. In the whole transformation experiments, 7 positive plants were obtained by PCR.