Resistance Analysis Of Transgenic Tobacco By2 Cell And Plant With Overexpression Of Coptis Japonica 4-hydroxyphenylpyruvate Dioxygenase Gene

HPPD is the key enzyme that catalyzes the formation of HGA from 4-HPP.It is very important for the biosynthesis of plastoquinone and vitamin E and plastoquinone is the key cofactor for biosynthesis of carotenoid. When it is inhibited, it will cause bleaching of the chlorophyll and finally the death of the plant. There is no report that natural HPPD has the resistance of herbicide.However, Professor Yuling Liang discovered that Cjhppd gene has the ability to resistant the herbicide. We transform this gene to tobacco BY2 cell and plant to take further explore in order to prove this gene can help resisting herbicide on the level of cell and plant. Therefore, it is very important for further study and application of this gene.We established the transformation and selection system of BY2 cell. We transformed the Cjhppd gene into BY2 cell by the method of Agrobacterium tumefaciens-mediated transformation and detected the transgenic cell by PCR. The results showed that all of the 12 transgenic cell line which was random selected were successfully transformed.Analysis of resistance of herbicide DTP showed that transgenic BY2 cell growed better than wild type in the presence of 30μmol/L DTP in fresh weight, color and other aspects. Transgenic BY2 cell contained more carotenoid and less MDA than that of wild type BY2 cell under DTP treated condition.The same thing happened on the plant. Analysis of resistance of herbicide DTP showed that transgenic plant growed better than wild type in the presence of DTP in color of leaves, plant height, root length and fresh weight. Transgenic plant contained higher chlorophyll a, chlorophyll b and carotenoid than that of wild type under DTP stress.It illustrated that overexpression of HPPD could enhance the resistance of herbicide in tobacco BY2 cell and plant.In addition, the results showed some resistance to NaCl of transgenic BY2 cell.Transgenic BY2 cell grows better than untransformed BY2 cell in the presence of NaCl.MDA detection showed that transgenic BY2 cell contained less MDA than that of untransformed BY2 cell. Under the stress of NaCl, transgenic one showed much better than that of wild type in germination of seeds, young plant growth situation, leaf differentiation and rooting of differentiation plant.