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【目的】以结瘤豆科植物紫花苜蓿根际土壤为研究材料,筛选具有ACC脱氨酶活力的氢氧化细菌,探索氢氧化细菌植物促生作用机制。【方法】利用持续通H2的气体循环培养体系、矿质盐固体培养基,分离、培养氢氧化细菌,观察菌株形态并测定生理生化特征;16SrDNA序列分析法构建系统发育树;采用薄层层析法筛选ACC脱氨酶阳性菌株,茚三酮显色法测定ACC脱氨酶活力。【结果】分离的37株细菌中有8株菌氧化氢和自养生长能力较强,初步确定为氢氧化细菌,从中筛选出1株ACC脱氨酶阳性菌株WMQ-7。菌株WMQ-7的形态特征、生理生化特征与恶臭假单胞菌(Pseudomonas putida)的特征基本一致;16SrDNA序列(GenBank登录号为EU807744)在系统发育树中与恶臭假单胞菌同属一个类群,序列同源性99%。鉴定菌株WMQ-7为恶臭假单胞菌,其ACC脱氨酶活力为0.671U/μg。【结论】采用气体循环培养体系分离氢氧化细菌,克服了传统配气法的局限。ACC脱氨酶阳性菌株的筛选,为深入研究氢氧化细菌作为植物根际促生菌的菌株特性和促生机制提供理论依据。
【Objective】 The rhizosphere soil of nodulated legume alfalfa was selected as the research material to screen the hydroxide bacteria with ACC deaminase activity to explore the mechanism of plant growth by hydrogen peroxide bacteria. 【Method】 The hydrogen peroxide bacteria were isolated and cultivated by the gas circulation culture system and the mineral salt solid medium with continuous H2 gas. Morphological and physiological and biochemical characteristics were observed and the phylogenetic tree was constructed by 16S rDNA sequence analysis. ACC deaminase-positive strains were screened, and the activity of ACC deaminase was determined by ninhydrin chromogenic assay. 【Result】 Among the 37 strains of isolated bacteria, 8 strains of bacteria had strong ability of hydrogen peroxide and autotrophic growth, and initially identified as hydrogen-oxidizing bacteria. One strain of ACC deaminase-positive strain WMQ-7 was screened out. The morphological, physiological and biochemical characteristics of the strain WMQ-7 were basically the same as those of Pseudomonas putida. The 16S rDNA sequence (GenBank accession number EU807744) belonged to the same group of Pseudomonas putida in phylogenetic tree, Sequence homology 99%. The strain WMQ-7 was identified as Pseudomonas putida and the activity of ACC deaminase was 0.671U / μg. 【Conclusion】 The gas-phase culture system was used to separate the hydrogen-oxidizing bacteria and overcome the limitations of the traditional gas distribution method. The screening of ACC deaminase-positive strains provides theoretical basis for further study on the strain characteristics and growth-promoting mechanism of hydrogen-oxidizing bacteria as plant growth-promoting rhizosphere bacteria.