摘要
【目的】对筛选到具有抑菌效果且产蛋白酶的菌株H进行分类地位的确定,同时对其产蛋白酶的条件进行优化,以期为其在植物病害防治上的应用奠定基础。【方法】通过生理生化、电子显微镜扫描及16sRNA测序相结合的方法明确菌株H的分类地位;通过单因素和正交试验对菌株H产蛋白酶条件进行优化,并检测其优化前后对病原真菌的抑菌效果。【结果】菌株H鉴定为高知芽孢杆菌(Cytobacillus kochii);菌株H产蛋白酶的优化条件为:pH8.0、葡萄糖20.0 g·L^(-1)、蛋白胨8.0 g·L^(-1)、MgSO_(4)1.0 g·L^(-1)、CaSO_(4)·2H_(2)O 0.1 g·L^(-1),在装有50 mL发酵培养基的250 mL三角瓶中接种2.5 mL10^(8) CFU·mL^(-1)的种子液,培养24 h后蛋白酶活力达到402.2 U·mL^(-1),较初始培养条件下的蛋白酶活力提高13.92倍;优化后菌株H对尖孢镰刀菌(Fusarium oxysporum)和辣椒疫霉病菌(Phytophthora capsici)的抑菌率分别为67.32%和44.87%,较优化前抑菌率提升值分别为9.15%和12.82%。【结论】明确菌株H为高知芽孢杆菌(C.kochii),优化了高知芽孢杆菌产蛋白酶条件及抑菌效果。
【Objective】Taxonomy of antimicrobial protease-producing Cytobacillus kochii H was studied for the development of a natural disease control agent on plants.【Method】Taxonomy of C.kochii H was determined by physiological,biochemical,electron microscopy scanning,and 16sRNA sequencing methods.A process of producing the antimicrobial protease secreted by the bacterium was optimized in single-factor and orthogonal experiments.The efficacy of the culture broth on 4 selected pathogenic fungi was applied in an in vitro test as the evaluation criterium.【Result】The optimized C.kochii H protease-producing process was conducted in a 250 mL triangular flask using 50 mL of pH 8.0 medium,which contained 20.0 g·L^(-1) of glucose,8.0 g·L^(-1) of peptone,1.0 g·L^(-1) of MgSO_(4),and 0.1g·L^(-1) of CaSO_(4)-2H_(2)O,inoculated with 2.5 mL of 10^(8) CFU·mL^(-1) C.kochii H fermentation liquid.After 24 h of incubation,the protease activity reached 402.2 U·mL^(-1),which was 13.92 times greater than it was prior to the optimization.The antimicrobial rates of the broth against Fusarium oxysporum and Phytophthora capsici were 67.32%and 44.87%,respectively,i.e.,9.15%and 12.82%,respectively,increases over those before the optimization.【Conclusion】The antimicrobial protease-producing C.kochii H was identified taxonomically.The culture process of the enzyme production was optimized to deliver significant in vitro inhibition rates on F.oxysporum and P.capsici.
作者
王学文
于存
潘洪祥
WANG Xuewen;YU Cun;PAN Hongxiang(Forestry College,Guizhou University,Guiyang,Guizhou 550025,China)
出处
《福建农业学报》
CAS
CSCD
北大核心
2023年第1期47-57,共11页
Fujian Journal of Agricultural Sciences
基金
国家自然科学基金项目(32160375)
黔科合平台人才项目([2018]5261)
关键词
高知芽孢杆菌
蛋白酶
产酶条件优化
病原菌抑菌率
Cytobacillus kochii
protease
optimized fermentation
antimicrobial rate on pathogens
