Application of a mineralizing carbon-fixing strain

Abstract

The invention discloses an application of a mineralized carbon-fixing strain, and the mineralized carbon-fixing strain is classified and named as sulfide mineral halomonas Halomonas sulfidearis. ZN2#, deposited in the General Microbiology Center of China Microbiological Culture Collection Committee, with the deposit number of CGMCC No.32033. The mineralized carbon fixation strain can metabolize urease and carbonic anhydrase at the same time. Combined with the microbial induced calcium carbonate precipitation technology, the two enzymes can undergo dual mineralization reactions, improve the crystal form of the mineralized product, and increase the mineralization intensity. At the same time, carbonic anhydrase can absorb carbon dioxide in the environment, achieve high-strength reinforcement of the soil body while reducing carbon dioxide emissions, thereby achieving the dual effects of soil biomineralization restoration and carbon fixation and emission reduction, and realizing the sustainable development of soil bioremediation and improvement technology.

Description

Technical Field

[0001] The present invention relates to the technical field of soil biomineralization restoration and carbon fixation and emission reduction, and specifically to a mineralized carbon fixation bacterial strain that simultaneously performs biomineralization restoration and carbon fixation and emission reduction on soil. Background Art

[0002] In recent years, with the advancement of global science and technology and the rapid development of the economy, all countries are facing various problems caused by the greenhouse effect. Carbon dioxide pollution is the main cause of the greenhouse effect. The greenhouse effect will also cause global warming, melting glaciers, and chaos in the food chain. Therefore, it is crucial to control carbon dioxide pollution and reduce carbon dioxide emissions. Microbial induced calcium carbonate precipitation technology, as a new soil remediation application technology, greatly reduces carbon dioxide emissions compared to traditional technologies. Microbial induced calcium carbonate precipitation (MICP) technology relies on the metabolic activities of microorganisms themselves. It utilizes the metabolism of high-yield urease alkaliphilic bacteria in nature, such as Bacillus pasteurianus, to produce urease, promote urea hydrolysis, and produce a large amount of CO3 2- It forms calcium carbonate precipitate with free calcium ions in the environment. Urease-producing bacteria adsorb on the surface of soil particles or sand and gravel particles. In the process of forming calcium carbonate, they cement the soil particles or sand and gravel particles, fill the pores in the soil, and thus achieve the purpose of soil strength and stiffness. However, in the process of forming calcium carbonate, some calcium carbonate with small particle size and unstable crystal form is often formed, which leads to low strength of the soil, and the respiration of microorganisms themselves will also increase carbon dioxide emissions. In recent years, a new technology uses the metabolic activities of microorganisms themselves to produce carbonic acid liver enzymes, such as Bacillus subtilis: by accelerating the CO2 hydration reaction, a large amount of CO3 2- It forms calcium carbonate precipitation with free calcium ions in the environment. However, in the process of forming calcium carbonate, a lot of calcium carbonate with small particle size and unstable crystal form will also be formed. Although it reduces the emission of carbon dioxide, it does not significantly improve the strength of the soil. However, there are no reports on the separation and screening of microorganisms that produce high urease and carbonic anhydrase from nature and the combination of MICP technology to achieve the goal of reducing carbon dioxide emissions while improving soil strength. Summary of the invention

[0003] The purpose of the present invention is to provide a highly efficient mineralization and carbon fixation strain and its application method, which can simultaneously produce urease and carbonic anhydrase for dual mineralization and soil carbon fixation and emission reduction to solve the problems raised in the above background technology. The purpose of the present invention is achieved through the following technical solutions:

[0004] A mineralizing carbon-fixing strain, named Halomonas sulfideminera ( Halomonassulfidaeris.) ZN2#, deposited in the General Microbiological Center of China Microbiological Culture Collection Administration, with the deposit number CGMCC No.32033 and the deposit date September 20, 2024. Its 16S rRNA gene sequence is shown in SEQ ID NO.1. Halomonassulfidaeris Esulfide 1 (or the same genus) has a homology of 99.08%.

[0005] The mineralized carbon-fixing strain is isolated from a salt lake. The strain belongs to Gram-positive bacteria, moderately halophilic bacteria and alkali-resistant bacteria, urease-producing mineralizing bacteria, and carbonic anhydrase-producing mineralizing bacteria. The optimal growth chloride concentration range of the mineralized carbon-fixing strain is 10%~15%; the chloride concentration range is 3%~20%; the sulfate concentration range is 0.4~1.5 mol / L; and the alkali resistance range is pH5.5~13. The strain appears as an irregular liquid water droplet on a solid culture medium, is white in color, and has a rod-shaped body with a length of 1μm~3μm, such as Figure 1 shown.

[0006] The logarithmic growth phase of the mineralizing carbon-fixing strain is 48 h, and the optimal culture conditions are: culture temperature 30 °C, culture speed 200 r / min, and culture time 48 h.

[0007] The separation and screening method of the mineralized carbon-fixing strain comprises the following steps:

[0008] Step 1: Take a water sample within the range of 0-10 cm below the surface of the salt lake, put it into a half-volume beef extract peptone culture medium after high-temperature sterilization, and shake it at a constant temperature of 25-32°C, a shaking frequency of 150-200 r / min, and a shaking time of 24 hours. After taking it out, let it stand for 12 hours; culture to obtain a bacterial enrichment solution;

[0009] Step 2: Collect the bacterial enrichment solution and prepare 10 -3 , 10 -4 , 10 -5 , 10 -6 , 10 -7 dilution of the dilution solution;

[0010] Step 3: The diluted solutions of different concentrations are respectively coated on a plate having a No. 1 solid culture medium on the surface and cultured at a constant temperature of 28°C to 32°C for 2 days to 3 days. The plate is sealed and placed in a constant temperature incubator for culture;

[0011] Step 4: Pick a single white colony on the solid culture medium and continue to inoculate it into the liquid culture medium for shaking culture. The shaking temperature is 25-32°C, the shaking frequency is 150-200 r / min, the pH is the natural pH of the culture medium, and the culture time is 24-48 hours;

[0012] Step 5: Take the bacterial solution to verify the production of urease;

[0013] Step 6: Take the bacterial solution to verify the production of carbonic anhydrase;

[0014] Step 7: Select a single colony that produces urease and carbonic anhydrase at the same time and continue to inoculate it into solid culture medium No. 1. After repeated streaking and purification, the obtained single colony is the mineralization carbon fixation strain;

[0015] The half-strength beef extract peptone culture medium comprises: 1-2 g beef extract, 5-10 g peptone, and 2-3 g NaCl per 1000 mL deionized water;

[0016] The solid culture medium comprises: solid culture medium No. 1: per 1000 mL of deionized water, 10-20 g of tryptone, 5-10 g of yeast extract, 100-150 g of NaCl, 5-15 mL of SL-6 trace element solution, and 10-15 g of agar;

[0017] Solid medium No. 2: 1 g glucose, 1 g casein, 1.5 g KH2PO4, 2 g Na2HPO4, 1 g MgSO4·7H2O, 0.012 g phenol red, 10-20 g urea, 10-15 g agar per 1000 mL deionized water;

[0018] The liquid culture medium comprises: 10-20 g of tryptone, 5-10 g of yeast extract, 100-150 g of NaCl, and 5-15 mL of SL-6 trace element solution per 1000 mL of deionized water.

[0019] Further, the urease production verification: different single colonies are inoculated into a No. 2 solid culture medium and cultured for 24 to 48 hours, and the color of the solid culture medium is observed. If the color of the culture medium changes from orange to magenta, it is judged that urease is produced by metabolism, and the single colony is a colony that produces urease;

[0020] The carbonic anhydrase production verification is as follows: different single colonies are placed in a liquid culture medium and cultured for 24 to 48 hours, then 5 to 10 ml of bacterial solution is inoculated into a calcium source solution, the mixed solution with white precipitation is allowed to stand for 24 hours, the supernatant is poured out, and the residual bacteria in the precipitation are washed with sterile distilled water, and the sediment sample is obtained by drying at 40 to 50°C for 12 to 24 hours; the sediment is detected with 1 mol / L dilute hydrochloric acid, and if a large number of bubbles are generated immediately after adding dilute hydrochloric acid to the sediment, the sediment is judged to be calcium carbonate precipitation, and the single colony is a colony that produces carbonic anhydrase; the calcium source solution only includes 0.5 to 1 mol / L dihydrate calcium chloride solution. The microscopic morphology of the sediment is observed and judged by a scanning electron microscope (SEM); the crystal structure of the sediment is analyzed by an X-ray diffractometer (XRD); and the chemical composition of the sediment is analyzed by an energy dispersive spectrometer (EDS).

[0021] The application of the mineralized carbon fixation strain in soil carbon fixation and emission reduction includes soil mineralization restoration and carbon fixation and emission reduction. The method includes the following steps:

[0022] Step 1): Prepare microbial solution and cementing solution for soil mineralization restoration and carbon fixation and emission reduction: The microbial solution is obtained by expanding the mineralization carbon fixation strain, and the OD of the solution is 600 The value is 1.0~2.0, and the urease activity is 4~6 mMurea / min; the composition of the cementing liquid is: urea 60~90 g / L, CaCl2·2H2O 147~221 g / L; the solvent of the cementing liquid is water;

[0023] Step 2): Perform in-situ microbial mixing to form a microbial mineralization layer: first, the microbial liquid and the cementing liquid are mixed in a volume ratio of 1:1-3 and stirred evenly to obtain a liquid-cementing liquid mixed liquid, and then the mixed liquid is mixed with soil in a mass ratio of 1:2-4 and stirred evenly, and finally the soil obtained by mixing is compacted and naturally air-dried until the strength reaches 1.5-5.0 MPa;

[0024] The mineralized carbon fixation strain produces urease and carbonic anhydrase through simultaneous metabolism, performs a dual mineralization reaction, improves the crystal form of the mineralized product, and effectively improves the solidification strength of the microbial mineralized soil. At the same time, through the action of carbonic anhydrase, carbon dioxide in the environment is captured, carbon emissions are reduced, and the dual effects of soil biomineralization restoration and carbon fixation and emission reduction are achieved.

[0025] The advantages and beneficial effects of the present invention are:

[0026] 1. The efficient mineralization and carbon fixation strain provided by the present invention is sulfide mineralizing bacteria Halomonassulfidaeris. ZN2# is a mineralizing strain that produces both high urease and high carbonic anhydrase.

[0027] 2. This highly efficient mineralized carbon fixation strain can capture carbon dioxide in the environment while inducing carbonate precipitation to solidify soil particles, achieving the dual effects of soil solidification and restoration and carbon fixation and emission reduction. It can be used as a new type of soil biological solidification and carbon fixation microbial strain, with great research and application value.

[0028] 3. The high-efficiency mineralization and carbon fixation strain is combined with microbial induced mineralization technology for microbial mineralization. Through the dual mineralization reaction, the crystal form of the mineralized product is improved, thereby effectively improving the strength of the soil consolidated by microbial mineralization. It can be used as a new type of high-strength mineralizing microorganism for soil biomineralization reinforcement, and has great application potential. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] The present invention will be further described below in conjunction with the accompanying drawings and embodiments.

[0030] Figure 1 The sulfide mineral salt monosaccharide provided in the embodiment of the present application Halomonassulfidaeris. ZN2# colony morphology and SEM scanning electron microscopy images;

[0031] Figure 2 The sulfide mineral salt monosaccharide provided in the embodiment of the present application Halomonassulfidaeris. Phylogenetic tree of ZN2#;

[0032] Figure 3 The sulfide mineral salt monosaccharide provided in the embodiment of the present application Halomonassulfidaeris. SEM, EDS and XRD spectra of ZN2# carbonic anhydrase-induced mineralization products; Figure 3 (a) is SEM, Figure 3 (b) for EDS, Figure 3 (c) is the XRD pattern;

[0033] Figure 4 The sulfide mineral salt monosaccharide provided in the embodiment of the present application Halomonassulfidaeris. SEM, EDS and XRD spectra of the mineralization products induced by the combined action of urease and carbonic anhydrase in ZN2#; Figure 4 (a) is SEM, Figure 4 (b) for EDS, Figure 4 (c) is the XRD pattern;

[0034] Figure 5 The sulfide mineral salt monosaccharide provided in the embodiment of the present application Halomonassulfidaeris. Growth curve of ZN2#;

[0035] Figure 6 The sulfide mineral salt monosaccharide provided in the embodiment of the present application Halomonassulfidaeris. Urease activity bar graph of ZN2#;

[0036] Figure 7 The sulfide mineral salt monosaccharide provided in the embodiment of the present application Halomonassulfidaeris. Carbonic anhydrase activity bar graph of ZN2#;

[0037] Figure 8 The sulfide mineral salt monosaccharide provided in the embodiment of the present application Halomonassulfidaeris. ZN2# captures the carbon dioxide concentration curve;

[0038] Fig. 9 This is the strength characteristic (UCS) curve of the standard sand mineralized and consolidated by the high-efficiency mineralized carbon-fixing strain provided in the examples of the present application. DETAILED DESCRIPTION

[0039] Example 1

[0040] A mineralizing carbon-fixing strain, the mineralizing carbon-fixing strain is classified and named as Sulfide Mineral Halomonas ( Halomonassulfidaeris. ) ZN2#, deposited in the General Microbiological Center of China Microbiological Culture Collection Committee, with the deposit number CGMCC No.32033 and the deposit date September 20, 2024. Address: No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing. Its 16S rRNA gene sequence is shown in SEQ ID NO.1. Halomonassulfidaeris Esulfide 1 (or the same genus) has a homology of 99.08%.

[0041] The mineralizing carbon-fixing strain was isolated from a salt lake. The strain is a Gram-positive bacterium, a moderately halophilic bacterium and a facultative alkaliphilic bacterium, a urease-producing mineralizing bacterium, and a carbonic anhydrase-producing mineralizing bacterium. The strain has an optimal growth chloride concentration range of 10% to 15%; a chloride concentration tolerance range of 3% to 20%; a sulfate concentration tolerance range of 0.4 to 1.5 mol / L; and an alkali tolerance range of pH 5.5 to 13. The strain appears as an irregular liquid droplet-like morphology on a solid culture medium, is white in color, and has a rod-shaped body with a length of 1μm to 3μm. Figure 1 shown.

[0042] In this embodiment, the separation and screening of the mineralized carbon fixation strain includes the following steps:

[0043] Step 1: Isolation and screening of microorganisms from salt lakes, specifically including the following steps:

[0044] The samples used in this experiment mainly come from salt lakes. Salt lake samples were taken from Yuncheng Salt Lake in Shanxi.

[0045] Step 1: Take a water sample within the range of 0-10 cm below the surface of the salt lake, put it into a half-volume beef extract peptone culture medium after high temperature sterilization, and shake it at a constant temperature of 28-32°C, a shaking frequency of 150-200 r / min, and a shaking time of 24 hours. After taking it out, let it stand for 12 hours; culture it to obtain a bacterial enrichment solution;

[0046] The half-dosage beef extract peptone culture medium comprises: 1-2 g beef extract, 5-10 g peptone, and 2-3 g NaCl per 1000 mL deionized water;

[0047] Step 2: Collect the bacterial enrichment solution and prepare 10 -3 , 10 -4 , 10 -5 , 10 -6 , 10 -7 dilution of the dilution solution;

[0048] Step 3: The diluted solutions of different concentrations are respectively coated on a plate having a No. 1 solid culture medium on the surface and cultured at a constant temperature of 28°C to 32°C for 2d to 3d, and the plate is sealed and placed in a constant temperature incubator for culture; No. 1 solid culture medium: per 1000mL of deionized water, including 10-20g of tryptone, 5-10g of yeast extract, 100-150g of NaCl, 5-15mL of SL-6 trace element solution, and 10-15g of agar;

[0049] Step 4: Pick a single white colony on the solid culture medium and continue to inoculate it into the liquid culture medium for shaking culture. The shaking temperature is 28-32°C, the shaking frequency is 150-200 r / min, the pH is the natural pH of the culture medium, and the culture time is 24-48 hours;

[0050] Wherein, the liquid culture medium is LB liquid culture medium: composed of 10-20 g tryptone, 5-10 g yeast extract, 100-150 g NaCl, and 5-15 mL SL-6 trace element solution;

[0051] Step 5: Verification of bacterial urease production,

[0052] The verification of the urease production property of the strain comprises the following steps:

[0053] a. Inoculate a single colony from any of the above-mentioned mineralizing carbon fixation strains during isolation into solid culture medium No. 2 and culture for 24h~48h

[0054] b. Observe the color of the solid culture medium. If the color of the culture medium changes from orange to magenta, it is judged that urease is produced by metabolism, and the single colony is a colony that produces urease;

[0055] Wherein, the second solid culture medium is a urea solid culture medium, which is composed of 1g glucose, 1g casein, 1.5g KH2PO4, 2g Na2HPO4, 1g MgSO4·7H2O, 0.012g phenol red, 10-20g urea, and 10-15g agar;

[0056] Step 6: Take the bacterial solution to conduct mineralization performance verification experiment.

[0057] The verification of the mineralization performance of the strain comprises the following steps:

[0058] a. Inoculate a single colony obtained during the isolation of any of the above-mentioned mineralized carbon fixation strains into a liquid culture medium and culture for 24 to 48 hours, then take 5 to 10 ml of the bacterial solution and mix it with a calcium source solution, and let the mixed solution with a white precipitate stand for 24 hours, wherein the calcium source solution only includes 0.5 to 1 mol / L calcium chloride dihydrate solution;

[0059] b. Pour off the supernatant, wash with sterile distilled water to remove the residual bacteria in the sediment, and dry at 40-50 °C for 12-24 h to obtain a sediment sample;

[0060] c. Use 1 mol / L dilute hydrochloric acid to detect the sediment. If a large number of bubbles are generated immediately after adding dilute hydrochloric acid to the sediment, it is judged that the sediment is calcium carbonate precipitation, and the single colony is a colony with mineralization performance;

[0061] d. Further, a scanning electron microscope (SEM) is used to observe and determine the microscopic morphology of the sediment; an X-ray diffractometer (XRD) is used to analyze the crystal structure of the sediment; and an energy dispersive spectrometer (EDS) is used to analyze the chemical composition of the sediment. The analysis results are as follows: Figure 3 , Figure 4 .

[0062] Figure 3 To characterize the mineralization products induced by carbonic anhydrase of the strain, Figure 3 (b) is the EDS spectrum, which shows that the elements that induce mineralization products are mainly C, O, and Ca. Figure 3 (a) is a SEM image, which shows that the induced mineralization product is in an unfixed spherical shape. Combined with the XRD spectrum, it is determined that the crystal form of the mineralization product induced by the carbonic anhydrase of the strain is mainly aragonite; Figure 4 To characterize the mineralization products induced by the combined action of urease and carbonic anhydrase of the strain, Figure 4 (b) is the EDS spectrum, which shows that the elements that induce mineralization products are mainly C, O, and Ca. Figure 4 (a) is a SEM image, which shows that the induced mineralization product is square in shape. Combined with the XRD spectrum, it is determined that the crystal form of the mineralization product induced by the combined action of strain urease and carbonic anhydrase is mainly calcite.

[0063] Step 7: Select a single colony that produces urease and carbonic anhydrase and continue to inoculate it into solid culture medium No. 1. After repeated streaking and purification culture for 4 to 6 times, the single colony obtained is the efficient mineralization and carbon fixation strain Sulfide Mineral Halomonas Halomonassulfidaeris. ZN2#;

[0064] The sulfide mineral salt mononas is in the form of irregular liquid water droplets on the solid culture medium, and the color is white. The SEM scanning electron microscope shows that the bacteria are rod-shaped, and the length is 1 μm to 3 μm. Figure 1 shown.

[0065] Step 2: Identification of bacterial species. The obtained strains were screened and the 16S rDNA sequence was determined, and compared with the known sequences in the gene bank (Gen Bank) established by the National Center for Biotechnology Information (NCBI, http: / / www.Ncbi.nlm.nih.gov) of the United States using the local alignment search tool (Blast). Based on the 16S rDNA sequencing results, a phylogenetic tree was constructed using MEGA4.1 software. This strain belongs to Gram-positive bacteria, and its 16S rDNA gene sequencing results are shown in SEQ ID NO.1. The phylogenetic tree constructed based on the gene sequencing results is shown in Figure 2 The bacteria screened were confirmed to be of the genus Halomonas and named Halomonas sulfide Halomonassulfidaeris. ZN2# was deposited in the General Microbiology Center of China Microorganism Culture Collection Administration on September 20, 2024, and the registration number of the collection center is CGMCC No.32033.

[0066] Step 3: Sulfide mineralizing bacteria Halomonassulfidaeris. Growth characteristics of ZN2#. Figure 5 The logarithmic growth period of ZN2# bacteria is 2 to 48 h, and it enters the stable period after 48 h.

[0067] Step 4: Sulfide mineralizing bacteria Halomonassulfidaeris. Urease activity of ZN2#. Figure 6 The urease activity of ZN2# bacteria ranged from 0 to 6 mM urea / min, which was a high-producing urease bacterium.

[0068] Step 5: Sulfide mineralizing bacteria Halomonassulfidaeris. Carbonic anhydrase activity of ZN2#. Figure 7 The carbonic anhydrase activity of ZN2# bacteria ranged from 0 to 0.5 U / mL, which was a high-producing carbonic anhydrase bacterium.

[0069] Step 6: Sulfide mineralizing bacteria Halomonassulfidaeris.The carbon dioxide capture characteristics of ZN2# are tested by titration: carbon dioxide hydration reaction generates carbonic acid, carbonic acid is partially ionized in water to generate hydrogen ions and bicarbonate ions, bicarbonate ions react with hydroxide ions under alkaline conditions to generate carbonate ions, and the amount of bicarbonate ions and carbonate ions is titrated to characterize the carbon dioxide capture characteristics. Figure 8 , ZN2# bacteria can capture 968ppm of carbon dioxide per liter of bacterial liquid.

[0070] Example 2

[0071] The application of the mineralized carbon-fixing strain in soil carbon fixation and emission reduction includes the following steps: This example uses the mineralized carbon-fixing strain isolation and screening method of Example 1 to isolate the strain.

[0072] Step 1): Prepare microbial solution and cementing solution for soil mineralization restoration and carbon fixation and emission reduction: The microbial solution is obtained by expanding the mineralization carbon fixation strain, and the OD of the solution is 600 The value is 1.8, the urease activity is 4-6 mMurea / min, and the carbonic anhydrase activity is 0.1-0.5 U / mL; the composition of the cementing liquid is: urea 60.06 g / L, CaCl2·2H2O 147.02 g / L; the solvent of the cementing liquid is water;

[0073] Step 2): The mixing method is used to prepare the microbial mineralization specimen, comprising the steps of: firstly, mixing the microbial liquid with the binder in a volume ratio of 1:1 and stirring evenly, then mixing the mixture of the microbial liquid and the binder with the standard sand in a mass ratio of 1:3.17 and stirring evenly, loading into a prefabricated mold and compacting; then placing in an air bath thermostat for curing at a constant temperature of 30°C for 5 days, and naturally air-drying for 14 days after the curing.

[0074] The mineralized carbon fixation strain produces urease and carbonic anhydrase through simultaneous metabolism, performs a dual mineralization reaction, improves the crystal form of the mineralized product, and effectively improves the solidification strength of the microbial mineralized soil. At the same time, through the action of carbonic anhydrase, carbon dioxide in the environment is captured, carbon emissions are reduced, and the dual effects of soil biomineralization restoration and carbon fixation and emission reduction are achieved. Figure 8 The sulfide mineralizing bacteria provided in the embodiments of the present application Halomonassulfidaeris . ZN2# captures carbon dioxide concentration curve, each liter of bacterial liquid can capture 968ppm of carbon dioxide; Fig. 9 The sulfide mineralizing bacteria provided in the embodiments of the present application Halomonassulfidaeris . The strength characteristic (UCS) curve of ZN2# mineralized consolidation standard sand shows that the solidification strength can reach 4.75MPa, which is 137.5% higher than the average strength of soil solidified by traditional mineralized strains (comparative example).

[0075] Example 3

[0076] The application of the mineralized carbon-fixing strain in soil carbon fixation and emission reduction includes the following steps: This example uses the mineralized carbon-fixing strain isolation and screening method of Example 1 to isolate the strain.

[0077] Step 1): Prepare microbial solution and cementing solution for soil mineralization restoration and carbon fixation and emission reduction: The microbial solution is obtained by expanding the mineralization carbon fixation strain, and the OD of the solution is 600 The value is 1.0, the urease activity is 4-6 mMurea / min, and the carbonic anhydrase activity is 0.1-0.5 U / mL; the composition of the cementing liquid is: urea 60.06 g / L, CaCl2·2H2O 147.02 g / L; the solvent of the cementing liquid is water;

[0078] Step 2): The microbial mineralization specimen is prepared by a mixing method, comprising the steps of: firstly, mixing the microbial liquid and the binder in a volume ratio of 1:1 and stirring evenly, then mixing the mixture of the microbial liquid and the binder with standard sand in a mass ratio of 1:3.17 and stirring evenly, loading into a prefabricated mold and compacting; then placing in an air bath thermostat for curing at a constant temperature of 30°C for 5 days, and naturally air-drying for 14 days after the curing.

[0079] The mineralized carbon fixation strain produces urease and carbonic anhydrase by simultaneous metabolism, performs a dual mineralization reaction, improves the crystal form of the mineralized product, and effectively improves the solidification strength of the microbial mineralized soil. At the same time, through the action of carbonic anhydrase, carbon dioxide in the environment is captured, carbon emissions are reduced, and the dual effects of biomineralization and restoration of soil and carbon fixation and emission reduction are achieved. The effect of this embodiment is slightly lower than that of Example 2, and the difference is not significant.

[0080] Example 4

[0081] The application of the mineralized carbon-fixing strain in soil carbon fixation and emission reduction includes the following steps: This example uses the mineralized carbon-fixing strain isolation and screening method of Example 1 to isolate the strain.

[0082] Step 1): Prepare microbial solution and cementing solution for soil mineralization restoration and carbon fixation and emission reduction: The microbial solution is obtained by expanding the mineralization carbon fixation strain, and the OD of the solution is 600 The value is 0.5, the urease activity is 4-6 mMurea / min, and the carbonic anhydrase activity is 0.1-0.5 U / mL; the composition of the cementing liquid is: urea 60.06 g / L, CaCl2·2H2O 147.02 g / L; the solvent of the cementing liquid is water;

[0083] Step 2): The microbial mineralization specimen is prepared by a mixing method, comprising the steps of: first, mixing the mineralized carbon-fixing bacteria solution and the cementing solution in a volume ratio of 1:1 and stirring evenly, then mixing the bacteria solution-cementing solution mixture with standard sand in a mass ratio of 1:3.17 and stirring evenly, loading into a prefabricated mold and compacting; then placing in an air bath thermostat for curing at a constant temperature of 30°C for 5 days, and naturally air-drying for 14 days after the curing.

[0084] The efficient mineralization carbon fixation strain produces urease and carbonic anhydrase by simultaneous metabolism, performs a dual mineralization reaction, improves the crystal form of the mineralized product, and effectively improves the solidification strength of the microbial mineralized soil. At the same time, through the action of carbonic anhydrase, carbon dioxide in the environment is captured, carbon emissions are reduced, and the dual effects of biomineralization restoration of soil and carbon fixation and emission reduction are achieved. The effect of this embodiment is slightly lower than that of Example 2.

[0085] Example 5

[0086] The application of the mineralized carbon-fixing strain in soil carbon fixation and emission reduction includes the following steps: This example uses the mineralized carbon-fixing strain isolation and screening method of Example 1 to isolate the strain.

[0087] Step 1): Prepare microbial solution and cementing solution for soil mineralization restoration and carbon fixation and emission reduction: The microbial solution is obtained by expanding the mineralization carbon fixation strain, and the OD of the solution is 600 The value is 0.5, the urease activity is 4-6 mMurea / min, and the carbonic anhydrase activity is 0.1-0.5 U / mL; the composition of the cementing liquid is: urea 90.09 g / L, CaCl2·2H2O 220.53 g / L; the solvent of the cementing liquid is water;

[0088] Step 2): The mixing method is used to prepare the microbial mineralization specimen, comprising the steps of: firstly, mixing the mineralized carbon-fixing bacteria solution and the cementing solution in a volume ratio of 1:2 and stirring evenly, then mixing the bacteria solution-cementing solution mixture with standard sand in a mass ratio of 1:2 and stirring evenly, loading into a prefabricated mold and compacting; then placing in an air bath thermostat for curing at a constant temperature of 30°C for 5 days, and naturally air-drying for 14 days after the curing.

[0089] The efficient mineralization carbon fixation strain produces urease and carbonic anhydrase by simultaneous metabolism, performs a dual mineralization reaction, improves the crystal form of the mineralized product, and effectively improves the solidification strength of the microbial mineralized soil. At the same time, through the action of carbonic anhydrase, carbon dioxide in the environment is captured, carbon emissions are reduced, and the dual effects of biomineralization restoration of soil and carbon fixation and emission reduction are achieved. The effect of this embodiment is slightly lower than that of Example 2.

[0090] Example 6

[0091] The application of the mineralized carbon-fixing strain in soil carbon fixation and emission reduction includes the following steps: This example uses the mineralized carbon-fixing strain isolation and screening method of Example 1 to isolate the strain.

[0092] Step 1): Prepare microbial solution and cementing solution for soil mineralization restoration and carbon fixation and emission reduction: The microbial solution is obtained by expanding the mineralization carbon fixation strain, and the OD of the solution is 600 The value is 0.5, the urease activity is 4-6 mMurea / min, and the carbonic anhydrase activity is 0.1-0.5 U / mL; the composition of the cementing liquid is: urea 90.09 g / L, CaCl2·2H2O 220.53 g / L; the solvent of the cementing liquid is water;

[0093] Step 2): The microbial mineralization specimen is prepared by a mixing method, comprising the steps of: firstly, mixing the mineralized carbon-fixing bacteria solution and the cementing solution in a volume ratio of 1:3 and stirring evenly, then mixing the bacteria solution-cementing solution mixture with standard sand in a mass ratio of 1:4 and stirring evenly, loading into a prefabricated mold and compacting; then placing in an air bath thermostat for curing at a constant temperature of 30°C for 5 days, and naturally air-drying for 14 days after the curing.

[0094] The efficient mineralization carbon fixation strain produces urease and carbonic anhydrase by simultaneous metabolism, performs a dual mineralization reaction, improves the crystal form of the mineralized product, and effectively improves the solidification strength of the microbial mineralized soil. At the same time, through the action of carbonic anhydrase, carbon dioxide in the environment is captured, carbon emissions are reduced, and the dual effects of biomineralization restoration of soil and carbon fixation and emission reduction are achieved. The effect of this embodiment is slightly lower than that of Example 2.

[0095] Comparative Example 1

[0096] Step 1): Prepare microbial solution and cementing solution for soil mineralization restoration and carbon fixation and emission reduction: The microbial solution is prepared by amplification of Bacillus pasteurianus (ATCC 11859), and the average OD of the solution is 600 The value is 0.7, and the average urease activity is 6.67 mM urea / min; the composition of the cementing liquid is: urea 60.06 g / L, CaCl2·2H2O 147.02 g / L; the solvent of the cementing liquid is water;

[0097] Step 2): The mixing method is used to prepare the microbial mineralization specimen, comprising the steps of: firstly, mixing the microbial liquid with the binder in a volume ratio of 1:1 and stirring evenly, then mixing the mixture of the microbial liquid and the binder with the standard sand in a mass ratio of 1:3.17 and stirring evenly, loading into a prefabricated mold and compacting; then placing in an air bath thermostat for curing at a constant temperature of 30°C for 5 days, and naturally air-drying for 14 days after the curing.

[0098] The average solidification strength of standard sand biomineralized by Bacillus pasteurianus (ATCC 11859) can reach 2.0 MPa.

[0099] Finally, it should be noted that the above is only used to illustrate the technical solution of the present invention rather than to limit it. Although the present invention has been described in detail with reference to the preferred arrangement scheme, a person skilled in the art should understand that the technical solution of the present invention may be modified or replaced by equivalents without departing from the spirit and scope of the technical solution of the present invention.

Claims

1. An application of a mineralizing carbon-fixing strain, characterized in that: The mineralized carbon-fixing strain is classified and named as Sulfide Mineral Halomonas ( Halomonas sulfidaeris ) ZN2#, deposited in the General Microbiology Center of China Microbiological Culture Collection Administration, with the deposit number CGMCC No.32033 and the deposit date September 20, 2024; the strain is used for simultaneous biomineralization restoration and carbon fixation and emission reduction of soil.

2. The use of the mineralizing carbon fixation strain according to claim 1, characterized in that: The following steps are involved: Step 1): Prepare microbial solution and cementing solution for soil mineralization restoration and carbon fixation and emission reduction: The microbial solution is prepared by expanding the mineralization carbon fixation strain, and the OD of the solution is 600 The value is 1.0~2.0, the urease activity is 4~6 mM urea / min, and the carbonic anhydrase activity is 0.1~0.5U / mL; the composition of the cementing liquid is: urea 60~90 g / L, CaCl2·2H2O147~221 g / L; the solvent of the cementing liquid is water; Step 2): Perform in-situ microbial mixing to form a microbial mineralization layer: mix the microbial liquid and the binder in a volume ratio of 1:1-3 and stir evenly to obtain a liquid-binder mixed liquid, then mix the mixed liquid with the soil in a mass ratio of 1:2-4 and stir evenly, and finally compact the soil obtained by mixing and naturally air-dry until the strength reaches 1.5-5.0 MPa.

3. The use of the mineralizing carbon fixation strain according to claim 2, characterized in that: The method for expanding the mineralized carbon-fixing strain in step 1) is: inoculating the mineralized carbon-fixing strain into a liquid culture medium and shaking the culture at a shaking temperature of 25-32°C, a shaking frequency of 150-200 r / min, a pH of the natural pH of the culture medium, and a culture time of 24-48 hours. The liquid culture medium is: for every 1000 mL of deionized water, add 10-20 g of tryptone, 5-10 g of yeast extract, 100-150 g of NaCl, and 5-15 mL of SL-6 trace element solution.

4. The use of the mineralizing carbon fixation strain according to claim 3, characterized in that: The method for expanding the mineralized carbon-fixing strain in step 1) is: inoculating the mineralized carbon-fixing strain into a liquid culture medium and shaking the culture at a shaking temperature of 30° C., a shaking frequency of 200 r / min, and a culture time of 48 hours.

5. The use of the mineralizing carbon fixation strain according to claim 1, characterized in that: The applicable scope of soil is clay soil, silt soil and sandy soil.

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