The aim of this study was to investigate inhibition spectrum,characteristics and antifungal mechanism of antagonistic protein of B.subtilis ZL2-70 which possesses strong antagonistic abilities to Verticillium dahilae Kleb.,and to offer a theoretical basis for biological control of cotton Verticillium wilt and development of transgenic resistant varieties.The inhibition spectrum of antifungal protein was studied and effects of heat,pH,proteinase,EDTA,inorganic ion on inhibiting activity were also investigated.Furthermore,cellulase and chitinase of the crude antifungal protein were determined and the inhibition to mycelium and spores was studied by different methods.Results indicated that,antagonistic protein of B.subtilis ZL2-70 against Verticillium dahilae has broad inhibition spectrum,which had strong inhibition activity to 21 different plant pathogens.It was relatively stable,could tolerate high temperature,EDTA and proteinases and keep activity in the range ofpH 5-10,but it was sensitive to Fe3+.The preliminary study on inhibition mechanism demonstrated that the antifungal protein could break down the mycelium and leak the protoplasm,turn the mycelium cells into bigger sacs and inhibit germination of conidia and the inhibition activity was consistent with cellulase and chitinase activity of antifungai protein.Therefore,the inhibition mechanism is deduced as mycelial growth and spore germination are inhibited by cell enzyme activity of the antagonistic protein.%[目的]旨在研究棉花黄萎病拮抗细菌枯草芽孢杆菌Bacillus subtilis ZL2-70抗菌蛋白的抑菌谱、理化性质以及抑菌机理,为棉花黄萎病生物农药的研制与开发奠定理论基础.[方法]采用抑菌圈法测定该抗菌蛋白的抑菌谱,以及温度、pH、无机离子、有机溶剂、蛋白酶对其稳定性的影响,并通过无菌小瓶等方法研究抗菌蛋白对大丽轮枝菌菌丝和孢子的抑制作用,测定了抗菌蛋白的纤维素酶和几丁质酶活力.[结果]棉花黄萎病拮抗细菌B.subtilis ZL2-70抗菌蛋白抑菌谱广,对19种植物病原菌表现出抑菌活性;该蛋白理化性质相对稳定,耐高温,在pH 5.0～10.0范围内抑菌活性较稳定,经EDTA处理后活性变化不大,对Fe3+敏感,对蛋白酶K、胃蛋白酶和胰蛋白酶不敏感;该抗菌蛋白对大丽轮枝菌的菌丝生长和孢子萌发有双重抑制作用,能使病原菌菌丝胞壁消融、原生质凝集渗漏、菌丝细胞泡囊化,并可抑制病菌孢子的萌发;抗菌蛋白具有纤维素酶和几丁质酶活性,且酶活力与抑菌活性有关.[结论]初步推断该抗菌蛋白通过多糖酶活力破坏大丽轮枝菌的细胞壁和抑制病原菌的孢子萌发而起到抑菌作用.
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