| Gladiolus root rot, also known as fusarium wilt and dry rot, caused by Fusarium oxyporum f. sp gladioli which could cause vascular disease has become one of the main diseases of gladiolus. Diseases affecting Gladiolus production include dry rot, brown rot, and plant wilting. Management of Gladiolus root rot is mainly through chemical soil fumigation and alternate husbandry. Chemical control is costly and often ineffective, and in time may lead to the occurrence of variants of the pathogen resistance even environmental and health hazardous. Therefore, attentions should rely more on pathogen resistant gladiolus varieties. However, Gladiolus is mainly breed by way of dividing cormlets in production, but the long-term vegetative propagation is apt to be infected with the virus and caused degradation. In vitro selection may represent an immediate and inexpensive way of increasing the resistance level of highly susceptible cultivars of gladiolus.The culture conditions for the production of crude toxin of Gladiolus root rot pathogen, selection of cell mutant lines resistant to root rot, changes of defense enzyme activity of gladiolus somatic mutant were systematically studied in this research. The purpose of this study was to establish selection system of cell mutant lines which resistance to Gladiolus root rot. It is important for exploring a new breeding way of gladiolus. The results as follows:1. The Gladiolus root rot pathogen, preliminarily identified as Fusarium oxysporum f. sp. gladioli, was successfully separated from the typical susceptible plants. PD liquid medium was suitable for Gladiolus root rot pathogen growth and Czapek liquid medium was the best for the crude toxin production. The optimum culture condition for the pathogen growth was at24-30℃, pH6-7, darkness, shaking conditions has no significant effect on growth of the pathogen. Crude toxin production of the pathogen was27℃, pH7.0, darkness continuous shaking. The crude toxin with strong thermal stability had obvious toxic effect on gladiolus seedling, causing seedlings wilting. The permeability of root cell membrane, root activity and content of malondialdehyde of two gladiolus cultivars reflected the toxic effect of crude toxin. The resistant level of cultivar ’wagon’to crude toxin was higher than cultivar’Super Rose’to some extent.2. One step and multiple steps screen methods were established to obtain the resistant cell mutant lines and the resistance identification were assessed. The resistance of callus to crude toxin could be improved selected the multiple steps. When the concentration of crude toxin was50%. Survial rate of two cultivar gladiolus callus by multiple steps screen method increased by228.33%and27.00%compared to that of the control. Survial rate of two cultivar gladiolus callus by multiple steps screen methods increased by19.00%and18.66%compared to that of the cne step screen method. The multiple steps screen method was to be optimal.3. When the concentration of crude toxin was50%. The survival number of adventitious buds which treated with EMS before selection was21. It was3times of the adventitious buds without the EMS treatment. The results also showed that the survival rate and growth rate of adventitious buds which treated with EMS before selection increased13.17%and24.16%compared to that of adventitious buds without the EMS treatment. The experimental results indicated that the frequency of plants resistant to crude toxin was increased with the EMS treatment of callus before selection.4. The results of resistance identification indicated that the resistant levels of the gladiolus mutants resistant to both crude toxin and pathogen were increased significantly compared with the control plantlets. And the gladiolus mutants were more tolerance to crude toxins than pathogen. Three regenerate plants of cultivar’wagon’and nine regenerate plants of cultivar’Super Rose’ were obtained in in vitro selection. Among them, eight of the resistant plants obtained after treatment with EMS.5. During the48h of treatment with the crude toxin, the change of PAL, PPO, and POD activities of the regenerate plants were similar. All of them increased rapidly at the beginning after treatment then decreased. Great difference of POD, PAL and PPO activities existed between resistant and susceptible cultivars after treatment as well:the higher the resistance was, the stronger their activity was on regenerate plants. PAL activities of the regenerate plants of two cultivar gladiolus were1.43and1.50times the control. PPO activities of the regenerate plants of two cultivar gladiolus were1.47and1.99times of the control, POD activities of the regenerate plants of two gladiolus cultivars were1.53and1.70times of the control. The results indicated that there was a significant positive correlation between the PAL, PPO, POD activities and the resistant level of the regenerate plants to crude toxin. |
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