| As a good ornamental plant with high merchantability, Rhododendron hybridum has been one of horticultural azalea cultivars with the best quality, the longest continuous blooming period and the best outlet for azaleas market. However, some problems exit in the production and application of azaleas, including that some of horticultural races or varieties are not easy to be identified and so the classification in term of the morphological characters only is often open to question, the high temperature in summer becomes a key restrictive factor in their popularization and application in China for the poor high temperature resistance of R.hybridum,and no report can be found about the application of some extra substance on R. hybridum,which have been found to induce the high temperature resistance of some plant. To resolve the problems mentioned above, in our research, several R. hybridum cultivars with popular gardening application are used to the researches about ISSR (inter-simple sequence repeat) polymorphic DNA markers, the effects of high temperature stress on growth morphology, some physio-biochemical indices on heat tolerance,photosynthesis and chlorophyll fluorescence, high temperature resistance induced by CaCl2 and SA, and the inventory survey of gardening application of R. hybridum. Main conclusions are as follows:1. By optimizing the concentrations of PCR components, the suitable ISSR-PCR system for R. hybridum is as follows: The 20 u L PCR volume includes 10 ng DNA templates, 0.25 mmol·L-1 dNTPs, 2.0 mmol·L-1 Mg2+, 1.0 U Taq DNA polymerase and 0.4 ? mol·L-1 primers. The optimal PCR process is 5 minutes at 94? for predenaturation, the follows by 40 cycles, each with 45 seconds at 94 ? for denaturation, 45 seconds at 54.4 ? for annealing, 1.5 minutes at 72 ? for extension, finally 7 minutes at 72 ? for extension and holding the samples at 4 ?. The ISSR-PCR of 11 cultivars of R. hybridum approves the suitability and stability of the system. The polymorphic bands can be accounted .for 70% with nice repeatability.2. ISSR was used to analyze the genetic diversity of 11 cultivars of R. hybridum. Thirteen primers produced a total of 124 sites, of which 106 sites were polymorphic. As analyzed by the software POPGENE 32,the genetic diversity of R. hybridum was high, with the effective alleles of 1.5564, Nei's gene diversity of 0.3207, Shannon's Information Index of 0.4742, and polymorphic sites of 85.48%. The genetic diversity of the cultivars is high. The ISSR results showed that the 11 cultivars could be identified by 8 polymorphic sites amplified by 6 ISSR primers. A computerized ISSR fingerprinting of the cultivars was drawn to facilitate the management of cultivars. The results confirmed that ISSR markers were available for the identification of R.hybridum.Cultivars.3. The research about the effects of high temperature stress on growth morphology of the four horticultural cultivars, Tiehong, Fenhong, Meihong and Puhong shows that, after four days in 38? or 43?,the both stresses make serious harm on the plants examined. However, some difference of growth morphology can be identified between the two temperatures and between the different cultivars. the injury made by the stress of 43 ? is faster and stronger than that made by 38?. And a small number of Tiehong,Puhong, and Meihong can recover after the elimination of stress, although none can recover which endured 43 ? stress.4. With the high temperature stress, the distinct changes can be distinguished on the relative conductivity,SOD activity, content of MDA, soluble protein, free proline, oxyradical and chlorophyll in the leaves of the four horticultural cultivars, Tiehong, Fenhong, Meihong and Puhong. The relative conductivity and the content of MDA and oxyradical will increase with the temperature and time of stress, nonetheless, the content of chlorophyll will decrease and the content of soluble protein will fall after rising. However, the content of proline will change indistinctively after four days stress, and the result suggests that the index is not suitable for the screening of heat-tolerant cultivars. Whether the SOD activity is a good identification index of heat resistance or not lies on the suitable temperature. Therefore, the heat tolerance of different cultivars can be evaluated nicely using some of the indices,such as the relative conductivity, the content of MDA, soluble protein and oxyradical. However, a single index may reflect differently to different cultivars, and a single cultivar may respond differently to different indices, and so the heat tolerance of a cultivar can not be estimated well by a single index.5. By the main component analysis from the seven individual indices, the contribution rate of soluble protein, MDA and relative conductivity can be of 61.6%, 23.2% and 12.1%, respectively. And the cumulative contribution rate can be of 96.8% so that the effects of the other indices can be ignored. Results of the subordinative function analysis show that the heat tolerance of the four cultivars is as follows:Tiehong>Puhong>Meihong>Fenhong. The results are consistent with the appearance of the growth morphology under the heat stress.6. The photosynthetic rate (Pn) of the four horticultural cultivars, Tiehong, Fenhong, Meihong and Puhong, decrease with the 38? or 43 ? heat stress, which suggests the stress makes the photoinhibition heavier, and the inhibition increases with temperature and time of stress. The result can also prove by the determination of the chlorophyll fluorescence parameters. The stomatal conductance (C) decreases and the concentration of internal leaf CO2 (intCO2) increases with the processing time, nonetheless, the transpiration rate (E) falls after rising. The results suggest that nomstomatal limitations are the key factors for the decrease of Pn during the 38? and 43 ? stress.7. The 38? and 43 ? stress can affect both of the photosynthetic electron transport process and quantasome, the values of Fv/Fm and ?PS? decrease distinctively. The results of recovery experiments, a small number of Tiehong, Puhong, and Meihong can recover after the elimination of stress, although none can recover which endured 43 ? stress, show that the heavy photoinhibitions have lead to non-recoverable harm or irreversive inactivation in the sruture of PS ?. The qP and NPQ also decrease distinctively with the heat stress. The results may suggest that the PS ? reaction centers are inactivated and the PQ reserves decrease rapidly by the heat stresss, and ultimately the leaves lose the ability of dissipating the overflowing excitation energy. The loss of the adaptability to self protection is consistent with the poor heat tolerance--one of the biological characteristics of R. hybridum.8. The relative conductivity and the content of soluble protein and MDA of Puhong and Meihong, can be improved in varying degrees after the plants were treated with 500mg·L-1?1000mg·L-1?1500mg·L-1 or 2000mg·L-1 CaCl2 solution. However, results of subordinative function analysis show that the best concentration is 1500mg·L-1.9. The relative conductivity and the content of soluble protein and MDA of Puhong and Meihong, can also be improved in varying degrees after the plants were treated with 50 mg·L-1, 100mg·L-1 or 150 mg·L-1 SA solution. However, results of subordinative function analysis show that the best concentration is 50mg ·L-1.10. By the on-the-spot investigation in Guangzhou, Hangzhou,Hongkong, Zhangping and Fuzhou city,China, the gardening application pattern of R. hybridum in the cities of China have changed from the pot culture at early time into pot flower, miniascape, exhibition, outdoor planting and specialized garden construction, and the plant arrangement have been in the present of diversity and ecological characteristics more and more. |
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