Study On The Tobacco And Potato Transformation With Arabidopsis Vacuole Membrane Na～+/H～+ Antiporter Gene(AtNHX1)

Na~+/H~+ antiporter in the large vacuole could separate Na~+ to vacuolar, it has the function of cytoplasm Na~+ detoxify and adjusting cell osmoregulation, and it is also necessary of halophytes and glycophytes salt-tolerance. This article transformed Na~+/H~+ antiporters from Arabidopsis-AtNHX1 to the mode plant tobacco and ascertained the gene's function. And then it was transformed to the goal plant potato and created salt tolerance potato germplasm, thus providing a useful model of breeding salt-tolerance potato cultivar. The major results as following:1.Optimization of tobacco's genetic transformation: (1)The Kanamycin selective concentration has been established: When the variety CV87 was the acceptor, the proper selective concentration of differentiation of tobacco leaf and the regeneration of roots were 50mg/L; When the variety T12 was the acceptor, the proper selective concentration of differentiation of tobacco leaf was 100mg/L and the regeneration of roots was 200mg/L; (2) Effects of cephalosporins on differentiation of tobacco leaf and inhibition of Agrobacterium tumefaciens: The effects of the six cephalosporins(Ceftriaxone sodium, Cefotaxime sodium, Cefoperzanoe sodium, Cefazolin sodium, Cefradine, Ceftazidime) in different concentrations were few on differentiation of tobacco leaf, the average differentiation rate was above 85% . 200mg/L Ceftriaxone sodium and Cefotaxime sodium had good effects in inhibiting Agrobacterium tumefaciens LBA4404, and they could eliminate the tobacco transgenic plant's bacterial cells after three generations(60d); (3)The genetic transformation conditions of tobacco were ascertained: leaf didn't pre-culture, OD600 of Agrobacterium tumefaciens was 0.3~0.5, infection Agrobacterium tumefaciens for 8~10min, co-culture 3 days;2. The transgenic plants of tobacco were obtained: (1) The transgenic plants which rooted in the Kanamycin medium were examined by distinctive PCR, and 1617bp positive signals were obtained, indication that AtNHX1 gene may be transformed into the genome of tobacco; (2)The manifest of transgenic tobacco's salt-tolerance: The salt-tolerance of transgenic plant was improved obviously, the transgenic plant could root in the 1.5%NaCl medium but the un-trangenic plant couldn't grow regularly;3. (1)The method of inducing potato microtuber has been established: liquid medium was best, there were 3 microtubers in every plant, the method was that plant growed in the conditions 25 days of MS~+3% sucrose, 25±1℃, 2500Lx light intensities, and then growed in MS~+8%sucrose~+5.00mg/L6-BA ~+0.15%C, after growing 2 days in the 2500Lx light...