9种豆科牧草根际促生菌筛选及特性研究

发布时间：2018-04-11 19:56

本文选题：豆科牧草 + 促生菌(PGPR)　；参考：《甘肃农业大学》2015年硕士论文

【摘要】：为收集豆科牧草优良菌种资源,为后续生物接种剂(菌肥)研制提供优质菌源,本研究利用钼蓝比色法和高效液相色谱法,以紫花苜蓿(Medicago sativa)、扁蓿豆(Trigonella ruthenica)、箭{H豌豆(Vicia sativa)、鹰嘴豆(Cicer arietinum)、小冠花(Coronilla varia)、鬼见锦鸡儿(Caragana jubata)、黄芪(Astragalus membranaceus)、毛苕子(Vicia villosa)、甘草(Glycyrrhiza uralensis)等9种豆科牧草为研究对象,从其根系及根际土壤分离、筛选根际促生菌(PGPR),测定菌株溶磷和分泌植物激素(生长素和赤霉素)特性,并利用乙炔还原法测定了扁蓿豆、箭{H豌豆、小冠花、毛苕子和黄芪5种植物根瘤的固氮酶活性,同时利用细菌形态、生理生化特性及16S rDNA序列分析对筛选出的优良促生菌株进行分类鉴定,主要结果如下:(1)从9种植物根际获得溶无机磷菌株分离物167个,根瘤菌5个,经过溶磷能力初步筛选,得到具有溶磷特性菌株66株,其溶磷量在1.3μg·mL-1~574.4μg·mL-1之间,溶磷量最大的是分离自鹰嘴豆根际的菌株PYRS6。各菌株溶磷能力差异显著(P㩳0.05),溶磷菌株的溶磷量与其培养液pH值之间具有一定的相关性,有5种植物根际溶磷菌株的溶磷量与其培养液pH值之间呈现极显著负相关性(P0.01)。与参比菌株Azotobacter beijerinckii相比,本次分离的菌株有75.8%的溶磷量优于参比菌株。(2)66株溶解无机磷菌株分泌植物激素的能力各不相同,其中,供试菌株分泌IAA量在2.09~43.3μg·mL-1之间,能力最强的菌株为PBRS5,分泌GA量在1.02~30.34μg·mL-1之间,分泌GA量最高的为PGRP9,并且有7株菌既具有分泌IAA能力,也兼具分泌GA的能力,其中以分离自鬼见锦鸡儿和毛苕子根际的菌株PGRP9、PGRS4和PMRS4表现较佳。(3)5种豆科牧草根瘤的固氮酶活性在1.21 umolC2h4/g·h~11.87 umolC2h4/g·h之间,根瘤固氮酶活性大小依次为:M-GL㧐B-GL㧐J-GL㧐X-GL㧐H-GL。(4)通过细菌形态观察、生理生化特定测定与分子生物学相结合的方法对15株优良PGPR(包括5株根瘤菌)进行分类鉴定,溶磷菌分属于6个属,分别为假单胞菌属、肠杆菌属、芽孢杆菌属、氮单胞菌属、固氮菌属以及克雷伯氏菌属,共8个种,即:Pseudomonas fluorescens、Enterobacter cloacae、bacillus subtilis、Azomonas agilis、Bacillus megaterium、Azotobacter chroococcum、Azotobacter vinelandii、Klebsiella oxytoca;4株根瘤菌分属于2个属,为慢生根瘤菌属和根瘤菌属。
[Abstract]:In this study, molybdenum blue colorimetry and high performance liquid chromatography (HPLC) were used to collect the fine strain resources of legume forage grass and to provide high quality bacteria source for the subsequent development of biological inoculants (bacterial fertilizer).Phytohormone and growth hormone (auxin and gibberellin) were determined, and the nitrogenase activities were determined by acetylene reduction method in five plant rhizobia, I. e., Astragalus root nodule, Astragalus membranaceus and Astragalus membranaceus, and the activity of nitrogenase was determined by acetylene reduction method, such as Astragalus membranaceus and Astragalus membranaceus.At the same time, the bacteria morphology, physiological and biochemical characteristics and 16s rDNA sequence analysis were used to classify and identify the excellent growth-promoting strains. The main results were as follows: 1) 167 isolates of soluble inorganic phosphorus and 5 rhizobia were obtained from the rhizosphere of 9 species of plants.After preliminary screening, 66 strains with the characteristic of phosphorus solubilization were obtained. The phosphorus soluble amount was between 1.3 渭 g mL-1~574.4 渭 g mL-1 and PYRS6, which was the highest in the rhizosphere of chickpea.There was a significant difference in phosphorus solubilization ability of each strain (P < 0.05). There was a certain correlation between phosphorus solubilization and pH value of the culture medium, and there was a very significant negative correlation between phosphorus solubilization and pH value in the culture solution of five plant rhizosphere strains.Compared with the reference strain Azotobacter beijerinckii, 75.8% of the strains were superior to the reference strain. The ability of dissolving inorganic phosphorus from 66 strains was different. Among them, the amount of IAA secreted by the tested strains was between 2.09 and 43.3 渭 g mL-1.The most potent strain was PBRS5, which secreted GA between 1.02g and 30.34 渭 g mL-1, and the highest amount of GA was PGRP9. There were 7 strains with both the ability to secrete IAA and the ability to secrete GA.Fifteen excellent PGPRs (including 5 rhizobia) were classified and identified by combining physiological and biochemical methods with molecular biology. Phosphorolytic bacteria belong to 6 genera, namely Pseudomonas, Enterobacter, Bacillus, Amonomonas.
【学位授予单位】：甘肃农业大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S54;S182

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