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Aspergillus niger strain with high yield of beta-glucosidase and application

A technology of glucosidase and Aspergillus niger strain, which is applied in the direction of glycosylase, enzyme, hydrolase, etc., can solve the problem of high cost of enzyme, utilization of biomass resources, low enzymatic efficiency of lignocellulosic hydrolase, cellulase system Problems such as low hydrolysis efficiency, to achieve the effect of improving efficiency and success rate, and enzyme system balance

Active Publication Date: 2021-09-17
TIANJIN INST OF IND BIOTECH CHINESE ACADEMY OF SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0002] At present, the enzymatic hydrolysis efficiency of lignocellulosic enzymes is low, and the high cost of enzymes is still one of the bottlenecks restricting the utilization of biomass resources. How to improve the enzymatic hydrolysis efficiency of cellulosic enzymes has always been the focus of research in the industry
However, the β-glucosidase content and enzyme activity in bacteria such as Trichoderma reesei, which are widely used in industrialization, are low, which has become one of the main limiting factors for the low hydrolysis efficiency of cellulase systems.

Method used

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  • Aspergillus niger strain with high yield of beta-glucosidase and application
  • Aspergillus niger strain with high yield of beta-glucosidase and application
  • Aspergillus niger strain with high yield of beta-glucosidase and application

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Experimental program
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Effect test

Embodiment 1

[0023] Embodiment 1. Obtain Aspergillus niger 60B-3DW strain by high-throughput screening method of β-glucosidase strain

[0024] Aspergillus niger ( Aspergillus niger ) 3.316 as the basis, the inventor through ARTP mutagenesis, with the help of droplet microfluidic high-throughput platform, by optimizing the conditions of Aspergillus niger droplet generation, sorting substrate and droplet size, the Aspergillus niger spores passed through the droplet package After burying and pre-cultivating for 16 hours, the droplet diameter was 80-90 µm, and the mycelium of Aspergillus niger did not grow too long to pierce the droplet, and at the same time, the mycelium of Aspergillus niger secreted enough β-glucosidase and fluorescent substrate Interaction, the level of enzyme activity in the droplet can be distinguished by the fluorescence value released by the substrate. It overcomes the drawbacks of the Aspergillus niger mycelial growth process that will cause the wrapped droplets to pu...

Embodiment 2

[0055] Embodiment two, the genetic stability test of Aspergillus niger 60B-3DW bacterial strain

[0056] In order to verify Aspergillus niger ( Aspergillus niger ) The genetic stability of the mutant strain 60B-3DW, after passing the mutant strain for 6 generations, it was fermented and cultured under the same fermentation conditions, and the change of its enzyme activity was compared. The average value of enzyme activity - the number of passages is plotted, and the results are shown in Figure 4 .

[0057] The enzyme activities of the mutant strain 60B-3DW in each generation were 88, 85.8, 87.6, 88.8, 88.5 and 86.3 IU / mL. It can be seen that the enzyme activities did not change much in these passages, which indicated that The mutant strain 60B-3DW has good genetic stability and can be used as an experimental strain for subsequent fermentation.

Embodiment 3

[0058] Embodiment three, the β-glucosidase of Aspergillus niger 60B-3DW fermentation and Trichoderma reesei cellulase hydrolysis biomass efficiency

[0059] Get Trichoderma reesei cellulase liquid and the Aspergillus niger β-glucosidase fermented liquid that obtains according to above-mentioned cultivation method according to different ratios (9.5:1, 8.5:1, 8.0:1, 7.5:1, 7:1 ) to form a compound cellulase preparation, and then measure the enzyme activity.

[0060] 1. Determination of filter paper enzyme activity

[0061] One enzyme activity unit (IU) is the amount of enzyme required to degrade and release 1 μmol of reducing sugar from the substrate (50 mg Whatman No.1 filter paper) per minute at 50°C and pH 4.8.

[0062] The assay of filter paper enzyme activity includes parallel and identical treatment of the following three parts of the experimental test tubes: blank control, glucose standard curve and fermentation broth detection.

[0063] ① Add 1.0 ml 0.05M, pH4.8 citric...

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Abstract

The invention belongs to the technical field of biomass material degradation and application of producing single-cell protein, and particularly discloses an aspergillus niger strain with high yield of beta-glucosidase and application. The preservation number of the aspergillus niger strain is CGMCC No. 22465, the activity of the beta-glucosidase produced by a fermentation method is up to 88IU / mL, a composite cellulase preparation is formed by simple compounding with a trichoderma reesei cellulase preparation, the enzyme system of the composite cellulase preparation is more balanced, the glucose release amount is increased by 35% or more under the condition of biomass high-solid enzymolysis, the cellulose conversion rate in the biomass material is greater than 90%, the application is completed in the straw single-cell protein, and the protein content in the straw single-cell protein product is greater than 25%.

Description

technical field [0001] The invention belongs to the technical field of biomass material degradation and its application in the production of single-cell protein, and relates to a strain of Aspergillus niger with high production of β-glucosidase and its application. Background technique [0002] At present, the low enzymatic efficiency of lignocellulosic hydrolytic enzymes and the high cost of enzymes are still one of the bottlenecks restricting the utilization of biomass resources. How to improve the efficiency of enzymatic hydrolysis of cellulase has always been the focus of research in the industry. The hydrolysis of lignocellulose is the result of the synergistic action of multi-enzyme systems, among which β-glucosidase is an important part of the cellulase system, which is responsible for the synergistic release of cellulosic oligosaccharides and Cellobiose is hydrolyzed to glucose. Therefore, by increasing the activity of β-glucosidase, not only the overall enzyme acti...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/14C12N9/42C12P19/14C12P19/02C12R1/685
CPCC12N1/14C12N9/2445C12N9/2437C12P19/14C12P19/02C12Y302/01021
Inventor 高乐吴信贾文娣
Owner TIANJIN INST OF IND BIOTECH CHINESE ACADEMY OF SCI
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