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Geobacillus thermodenitrificans HX-4 as well as method for producing cellulase and application of geobacillus thermodenitrificans HX-4

A technology of Geobacillus and HX-4, applied in the field of microorganisms, can solve the problems of decreased enzyme production capacity, poor cellulose effect, and low enzyme production by cellulose degrading bacteria, and achieve high cellulase activity and excellent Composting effect, effect of excellent cellulose degradation ability

Active Publication Date: 2022-05-13
HEBEI UNIVERSITY OF SCIENCE AND TECHNOLOGY
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  • Abstract
  • Description
  • Claims
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AI Technical Summary

Problems solved by technology

[0003] The current cellulose-degrading bacteria are not effective in decomposing cellulose, and the enzyme production of cellulose-degrading bacteria is relatively small, especially when the internal temperature of the compost increases, the enzyme-producing ability drops significantly, which cannot satisfy people's demand for high-efficiency cellulase strains. Requirements in ultra-high temperature environments

Method used

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  • Geobacillus thermodenitrificans HX-4 as well as method for producing cellulase and application of geobacillus thermodenitrificans HX-4
  • Geobacillus thermodenitrificans HX-4 as well as method for producing cellulase and application of geobacillus thermodenitrificans HX-4
  • Geobacillus thermodenitrificans HX-4 as well as method for producing cellulase and application of geobacillus thermodenitrificans HX-4

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] 1.1 Screening of strains

[0038] Sample collection: Samples were taken at a depth of 0.5m from the surface of the compost substrate in the aerobic compost substrate of the Food Waste Disposal Center in Shijiazhuang City.

[0039] Sample pretreatment: Take 10g of the above sample, add it to a sterile Erlenmeyer flask filled with 90mL of sterile water, seal it with a parafilm and shake it on a shaker at 150r / min for 220min, let it stand for 10min, take the suspension to obtain a bacterial suspension, and set aside .

[0040] Gradient dilution: Use sterile water to carry out 10-fold gradient dilution of the above bacterial suspension to obtain a concentration of 10 -2 、10 -3 、10 -4 and 10 -5 bacterial suspension.

[0041]Flat coating: in the ultra-clean operating table, take 10 -3 、10 -4 and 10 -5 0.1 mL of the bacterial suspension was added dropwise to the cellulose-Congo red medium for plate spreading and separation, and the inoculated medium was incubated upsid...

Embodiment 2

[0060] Using the Geobacillus thermodenitrifying HX-4 screened in Example 1 to produce cellulase and measure the enzyme activity.

[0061] 2.1 Method for producing cellulase

[0062] Step 1: inoculate Geobacillus thermodenitrifying HX-4 into LB medium with a pH of 7.0, culture in a shaker at 65°C and 190r / min for 24 hours, and prepare a seed solution.

[0063] Step 2: Add the seed solution to the fermentation medium with a 4% inoculation amount, and cultivate it on a shaker at 65°C, pH 7.0, and 190r / min for 72 hours to prepare a fermentation solution.

[0064] Step 3: Centrifuge the fermentation broth at 4°C and 4200 r / min for 10 minutes to remove bacteria, and take the supernatant as crude enzyme solution.

[0065] 2.2 Determination of cellulase activity

[0066] According to "QB 2583-2003", the enzyme activity of carboxymethyl cellulose was determined by the reducing sugar method, and the content of reducing sugar in the enzymatic solution was determined by the 3,5-dinitros...

Embodiment 3

[0072] The thermal denitrifying Geobacillus HX-4 screened in Example 1 was used to treat the sludge-rice husk powder mixed compost, and the change of the cellulose content in the mixed compost was determined.

[0073] Sludge-rice husk powder mixed compost: sludge and rice husk powder are mixed according to the weight of 4:1, the water content is controlled at about 65%, and 5% of the bacterial agent containing thermal denitrifying Geobacillus HX-4 is inoculated. Positive pressure aeration is adopted, and the aeration volume is 0.9m 3 / h, using intermittent aeration of "ventilate for 30 minutes, stop for 10 minutes". During the composting process, 0d, 3d, 5d, 8d, 14d, and 21d were turned over for sampling, and the total composting reaction time was 21d.

[0074] In this example, the American ANKOM A2000i automatic fiber measuring instrument was used to measure the ADF and ADL contents in the compost samples, and the cellulose content was measured by calculation (cellulose cont...

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Abstract

The invention relates to geobacillus thermodenitrificans HX-4 as well as a method for producing cellulase and application thereof. The preservation number of the geobacillus thermodenitrificans HX-4 is CGMCC (China General Microbiological Culture Collection Center) NO.24155; the geobacillus thermodenitrificans HX-4 can be used for preparing cellulase, the strain is inoculated into an LB culture medium to be cultured to prepare a seed solution, and the seed solution growing to the logarithmic phase is inoculated into a fermentation culture medium according to the inoculation volume of 3%-5% to be cultured to obtain fermentation liquor containing cellulase. The geobacillus thermodenitrificans HX-4 can normally grow under the condition of 55-70 DEG C, the activity of cellulase generated by the geobacillus thermodenitrificans HX-4 can reach 33.52 U / mL, after the geobacillus thermodenitrificans HX-4 is applied to sludge-rice hull powder mixed compost, the cellulose content of the geobacillus thermodenitrificans HX-4 is reduced by 26.81%, and the geobacillus thermodenitrificans HX-4 has great application value in the field of resource treatment and utilization of solid waste such as sludge and livestock and poultry manure.

Description

technical field [0001] The invention relates to the technical field of microorganisms, in particular to a thermodenitrifying geobacillus HX-4 and a method for producing cellulase and application thereof. Background technique [0002] Organic solid wastes such as rice husks, plant straws, sludge, and livestock and poultry manure contain a large amount of plant cellulose, which has a complex structure and is difficult to degrade, making it difficult to effectively realize resource utilization. At this stage, the efficient and stable resource utilization of plant cellulose at home and abroad basically relies on biological methods, using cellulase produced by microorganisms to degrade cellulose and make it into biological fertilizers. Aerobic composting is the mainstream treatment method to realize the resource utilization of organic solid waste. Compost maturity is a physiological and biochemical process dominated by microorganisms. The premise is to obtain microorganisms with ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12N9/42C12R1/01
CPCC12N9/2437Y02W30/40
Inventor 韩永辉滕志楠李清扬黄亚丽崔建升秦学安鸿雪李再兴周淋友王艳魁刘馨岳杨添琪吕建伟刘峰
Owner HEBEI UNIVERSITY OF SCIENCE AND TECHNOLOGY
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