一株兼具产IAA能力纤维素降解菌的筛选、鉴定及条件优化

doi:10.13560/j.cnki.biotech.bull.1985.2020-0454

摘要/Abstract

摘要：

从砂姜黑土中筛选兼具秸秆降解和植物促生能力的菌株,解决外源腐秆菌土壤存活状况差、促腐效果弱及作物生长不良等问题。结合羧甲基纤维素钠平板筛选、产CMC酶（内切-β-1,4-葡聚糖酶）和IAA（吲哚-3-乙酸）能力测定等评价菌株的秸秆降解和植物促生潜力;通过形态学、生理生化和16S rDNA序列分析进行菌株的种属鉴定;采用秸秆降解摇瓶试验和玉米盆栽试验对菌株实际促腐和促生功效进行测定;设置单因素试验探究菌株生长、产酶和产IAA的最适宜条件。筛出1株CMC酶活高、IAA产量大（20.15 mg/L）的菌株n3;经鉴定,n3为玉米固氮螺菌（Azospirillum zeae）;接种n3与对照相比,可将液态发酵下秸秆的促腐率显著提升54.71%,使土培条件下的玉米根长、平均直径和SPAD值分别提升18.3%、22.0%和5.24%（P<0.05）;n3生长、产酶和产IAA的最适氮源和装液量一致,分别为酵母粉和25 mL/250 mL,而产酶的最适pH为5.0,生长和产IAA的最适pH均为6.0。所筛玉米固氮螺菌（Azospirillum zeae）可用于生产多功能腐秆剂,加速砂姜黑土区还田秸秆腐解的同时促进作物的生长,开发利用的前景广阔。

关键词: 砂姜黑土, 玉米固氮螺菌, 秸秆促腐, 玉米促生, 条件优化

Abstract:

This study aims to screen strains with the ability of both degrading straw and promoting crop growth from Shajiang black soil for solving problems such as difficult survival status,weak rot-promoting effect and poor crop growth caused by exogenous rot straw. Screening with sodium carboxymethyl cellulose plate and combing with CMC enzyme（endo-1,4-β-D-glucanase）- and IAA（Indole-3-acetic acid）-producing capacity,the potential straw rot- and plant growth-promoting were evaluated. Species identification of strains were conducted via morphological,physiological and biochemical and 16S rDNA sequence analysis,then straw degradation shake flask and corn pot test were used to determine the actual rot- and growth-promoting effect of the strain,and a single factor test was set up to explore the optimal condition for growth and enzyme production of the strain. The screened strain n3 demonstrated the high activity of CMC enzyme（20.60 U/mL）and IAA synthesis（20.15 mg/L）,and was identified as Azospirillum zeae. The results showed that inoculation with n3 increased the rate of straw rot-promoting under liquid fermentation by 54.71%（P <0.01）compared with the control treatment;the root length,average diameter and SPAD value（Soil and plant analyzer development）of maize under soil culture conditions increased by 18.3%,22.0% and 5.24%,respectively（P<0.05）.The optimal nitrogen source and liquid volume of n3 growth,yielding CMC enzyme and IAA were the same as that of yeast powder and 25 mL/250mL,but the optimal pH was 5.0 for CMC enzyme production,and 6.0 for growth and IAA production. Conclusively,the screened A. zeae may be used to produce multi-functional straw-decomposing agent,accelerating the rot-decomposing of straw and promoting the growth of crops in the field,which has broad agricultural application prospect.

Key words: Shajiang black soil, Azospirillum zeae, straw rot-promoting, corn growth-promoting, optimization of conditions

吴婧, 聂彩娥, 朱媛媛, 黄薇, 马超, 姜瑛, 朱林, 郜红建. 一株兼具产IAA能力纤维素降解菌的筛选、鉴定及条件优化[J]. 生物技术通报, 2020, 36(12): 54-63.

WU Jing, NIE Cai-e, ZHU Yuan-yuan, HUANG Wei, MA Chao, JIANG Ying, ZHU Lin, GAO Hong-jian. Isolation,Identification of a Cellulose-degrading Bacterium with IAA-producing Ability and Optimization of Its Culture Conditions[J]. Biotechnology Bulletin, 2020, 36(12): 54-63.

图/表 6

图1 不同菌株产CMC酶（A）和IAA能力（B）不同字母表示差异显著（P<0.05）,下同

图2 菌株n3的菌落形态观察（A）和革兰氏染色（B）

表1 菌株n3的生理生化特征

项目	结果	项目	结果
明胶液化	-	柠檬酸盐利用	-
V-P试验	-	淀粉水解	-
M.R.反应	-	好氧性试验	好氧
接触酶试验	+	革兰氏染色	-

图3 基于n3和相关菌株的16S rDNA序列采用邻接法建立的系统发育树标尺代表每 100 个核苷中有 1 个核苷替代

表2 接种菌株n3对秸秆促腐和玉米生长的影响

处理	秸秆降解率/%	玉米生长
处理	秸秆降解率/%	SPAD值	平均直径/mm	根长/cm	根表面积/cm²	株高/cm	地上部鲜重/g
CK	9.76±0.90	40.2±1.62	0.41±0.04	23.5±2.29	111.5±23.0	51.8±1.25	3.91±0.42
n3	15.1±0.12**	42.3±0.82*	0.50±0.05*	27.8±5.02*	124.3±43.9	54.4±1.71	4.12±0.25

图4 不同培养条件对菌株n3生长状况、产CMC酶活力和IAA能力的影响 N1-N7：硝酸钾;硫酸铵;硝酸铵;酵母粉;谷氨酸;尿素;蛋白胨

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