The Responses Of Cucumber CsPLDα To NaCl Stress

Soil salinization is one of the major limited factors restricting the sustainable and efficient development of facility vegetables. Cucumber (Cucumis sativus L.) is the major vegetable cultivar in greenhouses in China, which is salt-sensitive and particularly sensitive to soil salinization. Phospholipase D (PLD) is a major hydrolase of phospholipid in the plant, the hydrolyzates including IP3, DAG, PA, and ethanolamine usually acts as a second messenger to participate in the signal transduction triggeing a series of reactions related to the resistance in the cell and involving in the growth, development as well as a variety of biotic and abiotic stresses. Thus, creating a new germplasm of CsPLDa-overexpression cucumber by genetic engineering and studing its function in NaCl stress acclimation could provide the theoretical basis for cucumber production in salinesoil. Based on previous studies, we obtained that transgenic tobaccos and cucumbers of CsPLDa-overexpression by Agrobacterium-mediated; through the study the expression pattens of CsPLDa and physiological characteristics by hydroponics to explore the response of CsPLDa to NaCl stress. The main results are shown as follows:1. VBlI121-CsPLDa (+) was transfered into the cucumber by Agrobacterium-mediated and5positive transgenic cucumbers were obtained after PCR detection. The transgenic cucumber, the To generation were generated by self-pollination, and then,5lines of the T1generation were obtaioned by PCR dection. we selected3positive transgenic T1generation cucumbers randomly, Real-time PCR detected that CsPLDa have been integrated into the genome of the cucumber and expressed.2. Meanwhiles we transferred pBI121-CsPLDa (+) into tobacco by Agrobacterium-mediated.90positive transgenic tobaccos were obtained by the PCR detection from98To generation tobaccos; we selected5positive transgenic tobaccos randomly, Real-time PCR detection showed that CsPLDa has been inserted into tobacco genome and expressed. Then the transgenic (T1-68,T1-71) and wild tobaccos were planted in the MS medium (with200mM NaCl), after9days, the tresult showed that the germination rates of transgenic tobaccos were95%and92.5%respectively, while the wild types were only about30%, which indicated that the tolerance to NaCl of transgenic tobaccos were significantly improved. 3. After200mM NaCl stress, the expression levels of CsNMAPK and CsPLDa and NO content were rapidly increased; and the expression levels of CsPLDa and NO content were increased up to the highest at2h; While the expression levels of CsNMAPK were significantly increased at0.5h. The above results showed that the expression of the CsPLDa lagged behind CsNMAPK in cucumber seedling leaves and it could stimulate the generation of NO.4. After200mM NaCl stress, the contents of H2O2and O2-in leaves of cucumber seedlings significantly increased. While the changes of the activities of antioxidant enzymes (SOD, POD, CAT and APX) were same, from0.5h to2h, decreased, then, increased gradually from4h to24h compared to Oh.5. After200mM NaCl stress from0.5h to2h, the contents of MDA, ABA and soluble protein in leaves of cucumber seedlings were significantly decreased compared to Oh; and then they both increased gradually, up to the highest at24h. The changes of GA and SA contents were declined, and then increased, later declined. SA content were reached the maximum after4h of treatment, while SA and GA contents were significantly lower after24h of treatment than0h.