Effects Of Excess Copper On Ascorbic Acid Releated Enzyme Activities And GalLDH Gene Expression In Non-Heading Chinese Cabbage

L-galactono-1, 4-lactone dehydrogenase (GalLDH) was a key enzyme that catalyzed the final step in the L-ascorbic acid synthesis pathway of plants. In our research, we studied the change of Vc anabolism enzymes in normal growth condition and copper treatment, and this gene could be used to increase ascorbate production, the nutritional value and stress tolerance through gene engineering technique.1. Ascorbic acid (AsA) contents, together with activities of L-galactono-1,4-lactone dehydrogenase (GalLDH), ascorbic acid oxidase (AAO), ascorbate peroxidase (APX), monodehydroascorbate reductase (MDAR) and dehydroascorbate reductase (DHAR) were measured during plant development in non-heading Chinese cabbage varieties ('Changzhouwutacai' ,'aijiaohuang' and 'Erqing'). The result showed that the content of AsA accumulated continually but first increased in the initial stage of plant development and then decreased, and the AsA content of 'Changzhouwutacai' has been higher than that of 'aijiaohuang' and 'Erqing'. Changes in GalLDH activity coincided with AsA accumulating rate during plant development, and GalLDH activity in 'Changzhouwutacai' was higher than that in other varieties. GalLDH activity was significantly positive correlated with AsA accumulation in three varieties. However, changes in activities of AAO and APX were in the opposite trend with AsA content, further more, AAO activity in 'Changzhouwutacai' was significantly lower than the other two varieties, APX activity but have no significant difference in varieties. MDAR and DHAR activity was determined mainly in the latter part of the growth. The results indicated that AsA content is regulated mainly by GalLDH activity in non-heading Chinese cabbage, AsA metabolizing enzymes—AAO, has also played an important role in the accumulation of AsA, and that the role of MDAR and DHAR for AsA accumulation is main in the latter part of the growth.2. Changes in ascorbic acid content and antioxidant enzyme activities in non-heading Chinese cabbage (Brassica chinensis) leaves of 'ChangzhouWutacai' and 'Erqing' exposed to excess copper (Cu) were investigated. Cu treatment reduced the fresh weight of shoot and root in both varieties, the reduction, however, was smaller in 'ChangzhouWutacai' compared to that in 'Erqing'. The higher copper tolerance of 'ChangzhouWutacai' may be due to its higher ascorbic acid content, which was mainly because of its lower activities of degrading enzymes, ascorbate oxydase (AAO) and ascorbate peroxidase (APX), as well as the increase of dehydroascorbate reductase (DHAR) activity after copper treatment compared with 'erqing'. Meanwhile, compared with 'erqing', the increase of catalase (CAT) and peroxidases (POD) activity in 'Wutacai' also played an important role in its copper tolerance.3. Changes in photosynthesis, the chlorophyll a fluorescence parameters, biomass, chlorophyll content, antioxidant and antioxidant enzyme activities, as well as reduced enzyme activities in non-heading Chinese cabbage (Brassica chinensis) leaves of 'ChangzhouWutacai' and 'Erqing' exposed to different copper (Cu) treatment were investigated. The result showed that there was difference between different varieties. The Cu tolerance in 'Changzhouwutacai' is higher than 'Erqing', the same with chlorophyll and AsA content. Antioxidant enzymes and AsA reduced enzymes have the same trends. However, it was better under 10μM copper than the control on biomass, chlorophyll content, Vc ontent, the chlorophyll a fluorescence parameters, antioxidant and AsA reduced enzymes.4. Changes in AsA content in different stage in non-heading Chinese cabbage (Brassica chinensis) leaves of 'ChangzhouWutacai' and 'Erqing' exposed to 75μM copper (Cu) treatment were investigated, and we also studied the expression of GalLDH gene by the method of Real-time RT-PCR. The result showed that the AsA content was increased significantly after Cu treatment and higher than the control. However, the expression of GalLDH gene increased remarkably with the change of Cu treatment, but reduced compared to the control. All of this indicated that the expression of GalLDH gene was not the main factor to the change of AsA content in non-heading Chinese cabbage, which was same with our other experiment. Moreover, the AsA content and expression of GalLDH gene in 'ChangzhouWutacai' were higher than in 'Erqing' .The results suggested that non-heading Chinese cabbage GalLDH was coded by more gene.