| Cucumber root-knot nematode is one of the important diseases in the process of cucumber production, causing serious loss and strong impact on the global agricultural economy. Though the chemical pesticides have significant preventative effects, they would be harmful to human and the environment. Therefore, biological control has been concerned for its advantages of non-toxicity and non-residual. Streptomyces roseoflavus Men-myco-93-63 is a biological agent that had evident antagonistic action against many plant pathogens, including root-knot nematode. In this study, cucumber seedling of Jinyou 1 treated with fermentation broth of Men-myco-93-63 was used to test the relative expression of WRKY genes in different tissue and different time by RT-PCR method, and proved up the control efficacy of Men-myco-93-63 tablets on cucumber root-knot nematode and its effect on soil microorganisms. Main results were as followed:1. The DNA was extracted from root-knot nematode and treated with the ITS universal primers for PCR amplification and sequencing, then using BLAST software to analysis sequence. The root-knot nematode collecting from Xinanhe of Dingzhou in Hebei was identified as Meloidogyne incognita.2. Six WRKY genes were isolated from cucumber seedling treated with aqueous solutions of Men-myco-93-63 using RT-PCR method, and the relative expression of 6 WRKY genes were tested in different tissues and different times. The results demonstrated that the relative expression of WRKY genes in different tissues was changed as the extension of treatment time. The expression of WRKY24 in root was higher than the treatment of root-knot nematode. In root, WRKY 17、WRKY23、WRKY24、WRKY48 WRKY49 and WRKY55 were up-regulated compared with the treatment of root-knot nematode, and the highest expression of WRKY17 and WRKY48 occurred 24 h after treatment, while WRKY 23 and WRKY49 were irregular. In stem, WRKY17、WRKY49、WRKY55 were up-regulated compared with the treatment of root-knot nematode, among which WRKY55 positive regulation was most significant; and the expression of WRKY23、WRKY48 were down-regulated. In leaf, WRKY17、WRKY48 WRKY49 and WRKY55 were higher than the treatment of root-knot nematode, and WRKY23 and WRKY49 up-regulated slightly. WRKY23 gene was regulated slightly by aqueous solutions of Men-myco-93-63, and its expression was close to the treatment of root-knot nematode. The regulatory effect the aqueous solutions of Men-myco-93-63 had on most WRKY transcription factors was higher in root that in stem and leaf. It can be speculated that the regulatory effect of WRKY transcription factors in root、stem and leaf caused by aqueous solutions of Men-myco-93-63 has some positive effects on the defense of root-knot nematode in cucumber, but the induction was different between different WRKY genes in different tissues.3. Greenhouse pot experiment and dilution-plate method were conducted to determine the control efficacy of the tablets on cucumber root-knot nematode and its effect on soil microorganisms. It showed that the control efficacy were 51.85% and 55.56% after treatment on cucumber by 1.0 g/plant and 1.5 g/plant, respectively, while that of the control agent-Paecilomyces lilacinus and solid fermentation of Men-myco-93-63 were 54.17% and 50.00%, respectively. Men-myco-93-63 tablets had marked effect on soil microorganisms after 60 days, the number of bacteria and actinomyces increased by 225.45% and 75.45% respectively, and the number of fungi reduced by 66.43% after treatment on cucumber by 1.0 g/plant. The total number of the 3 kinds of soil microorganisms increased by 195.35%, suggesting the tablets could significantly improve the structure of soil microorganisms. |
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