To Manipulate Cell Activity Of Cam Glutathione And Ascorbic Acid Metabolism In Transgenic Tobacco

Using two series of transgenic tobacco expressing cytosolic CaM as experimental materials, the effects of CaM activity on glutathione and ascorbate acid metabolism was studied.This research includes three parts:Part 1 CaM activity of 2 selected transgenic tobacco lines (H1-1C and H1-2C) were measured by phosphdiesterase. Then we studied the relationship between CaM activity and glutathione metabolism in transgenic tobacco. Tc-inducible transgenic tobacco expressing cytosolic CaM showed the higher contents of glutathione than those without Tc-induction and control (HO-2020). The more CaM activity of transgenic tobacco was increased, the more glutathione contents detected (such as H1-1C); On the contrary, after Tc-induction, the transgenic tobacco leaf disk were treated with CaM inhibitors, and then the glutathione contents were greatly decreased as the CaM activity did.The results above implied that CaM might play an important role in regulating glutathione metabolism in plants.Part 2 Based on the results of the Part 1, we studied the relationship between CaM activity and the most important restrict factors in the glutathione biosynthesis which is cysteine contents and γ-glutamylcysteine syhthetase(γ-ECS) activity. The results showed that the cysterine contents and γ-ECS activity of the transgenic tobacco were greatly improved after Tc-induction. When the CaM inhibitors were added to the transgenic tobacco leaf disk that after Tc-induction, the cysteine contents became higher and the γ-ECS activity obviously decreased than those without Tc-induction and control (HO-2020). In addition, we measured the activity of O-acetylserine(thiol)lyase (OAS-TL) which is the key enzyme of cysteine biosynthesis, and the enzyme showed the same change tendency as the cysteine did.Part 3 We studied the regulatory mechanism of CaM to control the GSG/GSSG and ASA/DHA ratio which represent the cell redox state. The results indicated that Tc-inducible transgenic tobacco showed the increased contents of ascobate acid than those without Tc-induction; On the contrary, when the CaM...