| Phalaenopsis spp is one of the most favorite flowers all over the world, which owns great market value. Tissue culture has been desired as the main way for mass propagation of Phalaenopsis. However, explant browning occurred during microprogation process is a vital problem, which could put explants to death and make serious economic loss in the following production. Many studies had been focused on the phenomenon of plant browning in order to explain the mechanisms of the browning and find the effective ways to solve this problem. But the present methods to control the plant browning have many limitations when they are applied to the tissue culture of Phalaenopsis. Thus, it is very necessary to study on the mechanisms of the tissue browning of Phalaenopsis to further understand the physiological basis on the process of the plant tissue browning.Two experiments had been carried out in this study. Firstly, the parameters of phenolic metabolism and reactive oxygen metabolism were measured in three varieties of Phalaenopsis B3,15 and F5 to study the physiological mechanisms of plant browning in genotype differences during the explants of three varieties inoculated in MS medium for 9 days. Secondly, the serious-browning varieties of Phalaenopsis B3 with low phenols content was used to study the effects and physiological basis of heat shock on the plant tissue browning. After heat shock treatment with 45℃for 18min and restoration for 5h, the leaf explants were sampled to measure the browning index, the parameters of phenolic metabolism and reactive oxygen metabolism every 3 days. The main results and conclusions are listed as follows:1. The total phenols content of the three varieties was positively correlated with their PAL,POD activitives. The variety B3 with lowest phenols content showed the most serious browning degree and highest PPO activity. However, the variety F5 with highest phenols content indicated the least browning degree and lowest PPO activity. The browning degree was not directly connected with the total phenols content. That the phenols in phalaenopsis were catalyzed by PPO may be the possible explanation of tissue browning occured. Therefore, the different PPO activity of the three varieties possibly was the one of main reasons that lead to browning differences.2. The MDA contents and the browning degrees were both gradually increasing during the tissue culture of Phalaenopsis,. Moreover, the browning degrees varied consistently MDA contents in the different varieties, indicating lipid peroxidation damage led to the tissue browning. SOD and APX as the main antioxidant enzymes participated in the scavenging of reactive oxygen species and alleviated the damage of membrance system. Thus, the different activities of antioxidant enzymes possibly were another reason about the genotype differences.3. It was shown that the explant browning of B3 was repressed significantly after heat shock treatment at 45℃for 18min and restoration for 5h. Phenol content and PAL,PPO activities on heat shock treatment were lower than control. However, the main reason of controlling browning by heat shock treatment could be dependent on the decrease of PPO activity of B3 containing low phenol.4. The activities of SOD,CAT,APX and GR were induced by heat shock treatment that were higher than the control at the initial period of tissue culture, especially the SOD and CAT activities of explant were rised quickly. It is very critical to protect the stability of the membrane system. Therefore, H2O2 and MDA were lower than the control on the whole culture days. Heat shock treatment could protect the stability of the membrane system by the enhancement of antioxidant system, thus the tissue browning of Phalaenopsis B3 was inhibited. |
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