具ACC脱氨酶活性大豆根瘤内生菌的筛选、抗性及促生作用

Screening, resistance and growth-promoting effect of endophytic bacteria with ACC deaminase activity isolated from soybean nodules

【目的】从大豆根瘤中筛选具ACC(1-氨基环丙烷-1-羧基)脱氨酶活性的内生细菌,对活性菌株的抗盐碱性、系统分类地位以及代表菌株的促生长作用进行研究,为发掘和应用抗逆、促生优良菌种资源提供理论基础。【方法】以ACC作为唯一氮源测定菌株产ACC脱氨酶特性,采用标准曲线法测定α-丁酮酸含量,比色法定量测定ACC脱氨酶活力,固体平板筛选法对活性菌株进行抗性分析,通过菌体形态及生理生化特性测定、16S rRNA基因序列同源性分析鉴定菌株分类地位,采用盆栽试验验证代表菌株的促生作用。【结果】从河南省13个市(地区)36个点采集的大豆根瘤中筛选出8株ACC脱氨酶内生细菌,其中菌株DD132的酶活性最高(15.712 U/mg)。筛选菌株可耐受4%–6%NaCl,其中菌株DD165、DD132可耐受9%NaCl盐浓度。在pH 11时5株(DD14、DD132、DD67、DD141、DD131)生长良好,说明这些菌株有较强耐碱性。8株产ACC脱氨酶菌株分属于4属,即芽孢杆菌属(Bacillus)、肠杆菌属(Enterobacter)、寡养单胞菌属(Stenotrophomonas)和泛菌属(Pantoea)。接种试验表明内生菌DD132对小麦幼苗生长具有明显促生长作用。【结论】大豆根瘤内具ACC脱氨酶高活性菌株有较强耐盐碱性,其中菌株DD132对小麦幼苗生长有明显促生长作用。为发掘和应用抗逆、促生的优良菌种资源提供理论基础。

[Objective] We screened endophytic bacteria containing ACC（1-aminocyclopropane-1-carboxylate）deaminase from soybean nodules, and evaluated salt-alkaline resistance, phylogenetic status and the growthpromoting of representative strain. [Methods] The features of strains producing ACC deaminase were determined by using the ACC as a sole nitrogen source, adopting standard curve method, colorimetric method, solid medium screening method, bacterial morphology, physio-biochemical characteristics, similarity analysis of 16 S rRNA gene,inoculation tests. [Results] Eight endophytic bacteria containing ACC deaminase were screened from soybean nodules collected from 36 points of 13 cities（regions）in Henan province. Enzymaticactivity of DD132 was the highest（15.712 U/mg）. Screened strain tolerated to medium of 4%–6% Na Cl concentration. Among of them, DD165 and DD132 could tolerate 9% NaCl concentration. Five bacteria growing well under pH 11, showing that these strains had stronger alkali resistance. Eight strains containing ACC deaminase activity were affiliated to four genera： Bacillus,Enterobacter, Stenotrophomonas and Pantoea. Inoculation test showed that DD132 had a significant growthpromoting effect on wheat seedlings. [Conclusions] Endophytic bacteria containing high ACC deaminase activity from soybean nodules have stronger salt-alkaline resistance. DD132 has obvious growth-promoting effect on wheat seedlings.