抗水稻细菌性条斑病芽孢杆菌菌株的选育研究

发布时间：2018-05-28 04:07

  本文选题：解淀粉芽孢杆菌 + 细菌性条斑病　； 参考：《广西大学》2017年硕士论文

【摘要】：解淀粉芽孢杆菌物种丰富,广泛分布在土壤和植物表面。近年来随着解淀粉芽孢杆菌具有抑制多种植物病原菌能力的发现,使其成为了具有生物农药发展潜力的微生物。从湖南、广西等地水稻田采取土壤样品,经分离、纯化得到9763株芽孢杆菌,采用牛津杯法测定分离菌株对水稻细菌性条斑病菌的拮抗能力,筛选得到具有拮抗能力的芽孢杆菌310株,经过多次重复筛选,最终得到拮抗能力强且稳定的菌株JF48,其对水稻细菌性条斑病的抑菌圈直径达37±3 mm。通过对该菌株的形态、生理生化以及16SrDNA序列进行分析,鉴定该菌株为解淀粉芽孢杆菌(Bacillus amyloliquefaciens)。通过摇瓶培养试验,研究了不同碳源、氮源、无机盐种类、摇瓶装液量、转速、起始pH、发酵培养温度、接种量对解淀粉芽胞杆菌JF48菌株芽孢产量及芽孢转化率的影响。然后通过Plackett-Burman试验从单因素试验得到的最佳因素中筛选出三个显著影响因子,分别为玉米粉、豆粕粉、发酵培养温度。最后,经响应面法优化后的最佳发酵培养条件为:玉米粉14.69g/L、豆粕粉 9.54g/L、CaC122g/L、NaCl5g/L、MgS04lg/L、装液量50mL/250mL、转速为200rpm、初始pH7.5、温度38℃、接种量1%。在此发酵培养条件下,芽孢产量达到3.6×109cffu/mL,与优化前相比提高了105 倍。通过单因素试验,确定解淀粉芽孢杆菌JF48菌株可湿性粉剂的最佳配方如下:载体硅藻土 10%、助剂聚乙二醇(PEG-8000)10%、稳定剂K2HPO42%、紫外保护剂抗坏血酸(VC)0.1%。使用该配方制成的解淀粉芽孢杆菌JF48菌株的可湿性粉剂芽孢含量为5.6×109cfu/g,悬浮率为76%,湿润时间为56s。使用JF48菌株培养液进行盆栽防治试验,试验组的水稻发病等级均明显低于对照组。这表明解淀粉芽孢杆菌JF48菌株对水稻细菌性条斑病具有明显的防治效果。
[Abstract]:Bacillus amylolyticus is abundant in species and widely distributed on soil and plant surfaces. In recent years, with the discovery of Bacillus amylolyticus which has the ability to inhibit many plant pathogens, it has become a potential microorganism with biological pesticide development. 9763 strains of Bacillus spp were isolated from paddy fields of Hunan and Guangxi provinces. The antagonistic ability of the isolated strains against bacterial spot bacteria of rice was determined by Oxford Cup method. 310 strains of Bacillus with antagonistic ability were screened. After repeated screening for many times, the strain JF48 with strong antagonistic ability and stability was obtained. The diameter of inhibition circle against bacterial streak disease of rice was 37 卤3 mm. The strain was identified as Bacillus amylolique faciensus by analyzing its morphology, physiology and biochemistry and 16SrDNA sequence. The effects of different carbon sources, nitrogen sources, inorganic salt species, liquid volume, rotation speed, initial pH, fermentation temperature and inoculation amount on spores yield and spores conversion of Bacillus amylolyticus JF48 strain were studied by shaking flask culture experiment. Then through Plackett-Burman test, three significant influencing factors were screened out from the best factors obtained by single factor test, which were corn flour, soybean meal powder and fermentation temperature. Finally, the optimum fermentation conditions were as follows: corn flour 14.69 g / L, soybean meal 9.54 g / L, CaC122g / L, NaCl5g / L, MgS04lg / L, 50 mL / L, rotational speed 200rpm, initial pH 7.5, temperature 38 鈩，

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