四株PGPR菌株混菌发酵体系的构建及促生效应评价

doi:10.13560/j.cnki.biotech.bull.1985.2020-0989

摘要/Abstract

摘要：

构建由4株PGPR菌株组成的混菌发酵体系并进行辣椒接种,为微生物复合菌剂的研制及开发奠定基础。通过单因素、正交实验及响应面法,优化4株菌混菌发酵的培养基和发酵条件,盆栽实验评价复合菌液对辣椒的促生效应。混菌发酵体系的最适碳氮源及比例为蔗糖∶酵母浸出物（5∶1）,最适发酵条件为：温度29.7℃、pH 7.05、接种量3.19%、装液量为50 mL/250 mL;该条件下制备的复合菌液活菌数为2.94×10¹⁴CFU/mL,是单菌混合菌液的5.13倍。盆栽实验显示复合菌液显著提高了辣椒幼苗的株高、根长、鲜重及叶绿素含量,较4株菌混合菌液接种分别提高了8.01%、50.22%、43.66% 和 6.94%。利用混菌发酵体系制备的复合菌剂有更高的活菌数,可更好地发挥对辣椒的促生效应,在PGPR复合菌剂开发中潜力明显。

关键词: 混菌发酵, 发酵条件优化, 复合菌剂, 辣椒促生

Abstract:

This objective of this work is to construct a mixed fermentation system from 4 plant growth-promoting rhizobacteria（PGPR）strains and to inoculate pepper for laying a foundation for the research and development of microbial compound agents. The medium and fermentation conditions of the mixed fermentation system were optimized by single factor experiment,orthogonal experiment and response surface method. And experiment of planting pepper in pot was conducted to evaluate the growth-promoting effect of compound broth. In mixed fermentation system,the optimal carbon source was sucrose,the optimal nitrogen source was yeast extract,and the optimal carbon-nitrogen ratio was 5∶1. The optimal fermentation conditions were as follows,temperature 29.7℃,pH 7.05,inoculation proportion 3.19%,and the medium volume 50 mL/250 mL. Under these culture conditions,the viable bacterial count in the mixed fermentation culture was 2.94×10¹⁴CFU/mL,which was about 5.13 times compared with single strain fermentation. The growth and physiological indexes of pepper seedlings were significantly improved after irrigating with the mixed fermentation broth,i.e.,the plant height,root length,fresh weight,and chlorophyll content increased by 8.01%,50.22%,43.66% and 6.94% compared with the single strain fermentation compound. In Conclusion,the compound agents prepared by the mixed fermentation system of 4 PGPR strains may have higher viable microbial counts,and play better growth-promoting effects on pepper,and have obvious application potential in the development of PGPR compound agent.

Key words: mixed fermentation, fermentation optimization, compound agent, growth-promoting effect of pepper

周静, 黄文茂, 秦利军, 韩丽珍. 四株PGPR菌株混菌发酵体系的构建及促生效应评价[J]. 生物技术通报, 2021, 37(4): 116-126.

ZHOU Jing, HUANG Wen-mao, QIN Li-jun, HAN Li-zhen. Construction of Mixed Fermentation System of Four PGPR Strains and Evaluation of Its Promoting Effect[J]. Biotechnology Bulletin, 2021, 37(4): 116-126.

图/表 13

参考文献 28

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碳源 Carbon source	碳源添加量Dosage/（g·L^-1）	氮源 Nitrogen source	氮源添加量Dosage/（g·L^-1）
果糖 Fructose	20.0	牛肉膏 Beef extract	12.08
蔗糖 Sucrose	19.0	酵母浸出物 Yeast extract	12.53
乳糖 Lactose	19.0	硫酸铵 Ammonium sulfate	6.84
麦芽糖 Maltose	19.0	硝酸钠 Sodium nitrate	8.79

碳源 Carbon source	碳源添加量Dosage/（g·L^-1）	氮源 Nitrogen source	氮源添加量Dosage/（g·L^-1）
果糖 Fructose	20.0	牛肉膏 Beef extract	12.08
蔗糖 Sucrose	19.0	酵母浸出物 Yeast extract	12.53
乳糖 Lactose	19.0	硫酸铵 Ammonium sulfate	6.84
麦芽糖 Maltose	19.0	硝酸钠 Sodium nitrate	8.79

水平 Level	因素Factors
水平 Level	A碳源 Carbon source	B氮源Nitrogen source	C碳氮比 Carbon/Nitrogen
1	葡萄糖Glucose	蛋白胨Peptone	1∶1
2	蔗糖Sucrose	牛肉膏 Beef extract	5∶1
3	乳糖Lactose	酵母浸出物Yeast extract	10∶1
4	麦芽糖Maltose	硫酸铵Ammonium sulfate	15∶1

水平 Level	因素Factors
水平 Level	A碳源 Carbon source	B氮源Nitrogen source	C碳氮比 Carbon/Nitrogen
1	葡萄糖Glucose	蛋白胨Peptone	1∶1
2	蔗糖Sucrose	牛肉膏 Beef extract	5∶1
3	乳糖Lactose	酵母浸出物Yeast extract	10∶1
4	麦芽糖Maltose	硫酸铵Ammonium sulfate	15∶1

试验号 Experiment No	A碳源 Carbon source	B氮源 Nitrogen source	C碳氮比 Carbon-Nitrogen ratio	D空白列 Blank	OD₆₀₀值 OD₆₀₀ value
1	1（葡萄糖Glucose）	1（蛋白胨Peptone）	1（1∶1）	1	0.741
2	1	2（牛肉膏Beef extract）	3（10∶1）	2	1.022
3	1	3（酵母浸出物Yeast extract）	4（15∶1）	3	1.747
4	1	4（硫酸铵Ammonium sulfate）	2（5∶1）	4	0.428
5	2（蔗糖Surcose）	1	2	4	1.428
6	2	2	4	3	1.619
7	2	3	3	2	1.805
8	2	4	1	1	0.228
9	3（乳糖Lactose）	1	3	2	1.463
10	3	2	1	1	1.509
11	3	3	2	4	1.628
12	3	4	4	3	0.154
13	4（麦芽糖Maltose）	1	4	3	1.313
14	4	2	2	4	1.590
15	4	3	1	1	1.607
16	4	4	3	2	0.250
K1	3.938	4.946	4.085	4.085
K2	5.080	5.740	5.075	4.540
K3	4.755	6.787	4.540	4.833
K4	4.760	1.060	4.833	5.075
k1	0.985	1.237	1.021	1.021
k2	1.270	1.435	1.269	1.135
k3	1.189	1.697	1.135	1.208
k4	1.190	0.265	1.208	1.269
R	1.142	5.727	0.990	0.990
因素主次Order of factors	B﹥A﹥C
最优组合Optimal combination	A₂B₃C₂