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

摘要:

[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。

Abstract:

[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 16S 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% NaCl 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.