Preparation method of biochar-loaded synthetic microbial agent and application of biochar-loaded synthetic microbial agent in plant growth promotion and soil remediation

Abstract

The invention discloses a preparation method of a biochar-loaded synthetic microbial agent and application of the biochar-loaded synthetic microbial agent in plant growth promotion and soil remediation, and relates to the technical field of plant growth promotion and soil remediation. The invention aims to solve the problems that the existing single degradation strain does not have the broad-spectrum PAEs degradation capability and the degradation effect of the single degradation strain on PAEs in indigenous bacterial communities is unstable. The synthesized microbial agent constructed by the invention shows more excellent performance on degradation and utilization of DBP, and the 48-hour degradation rate of the synthesized microbial agent on DBP is greater than the degradation rate of the synthesized microbial agent when only DNB-S1 degrading bacteria exist. The pores of the biochar provide a suitable environment for the habitat of microorganisms, so that the damage of high-concentration DBP to bacteria is avoided, the DBP adsorbed on the biochar is promoted to be utilized by DNB-S1, and the chemotaxis of bacteria is facilitated. The invention can obtain the preparation method of the biochar-loaded synthetic microbial agent and the application of the biochar-loaded synthetic microbial agent in plant growth promotion and soil remediation.

Description

Technical Field

[0001] The present invention relates to the technical field of plant growth promotion and soil remediation, and in particular to a method for preparing a biochar-loaded synthetic microbial agent and its application in plant growth promotion and soil remediation. Background Art

[0002] Phthalates (PAEs) are a class of compounds widely used as industrial plasticizers. As a plasticizer, no suitable substitute has been found so far. It is expected that the production and consumption of PAEs will remain high in the future. With the use of production materials such as agricultural films, greenhouses, pesticides and fertilizers, di(2-ethylhexyl) phthalate (DEHP) and dibutyl phthalate (DBP) are often widely detected in soil. As a source and sink of pollutants, the distribution of PAEs in soil is affected by factors such as human behavior, soil type and the properties of PAEs. The degree of degradation of PAEs by microorganisms depends on the structure and properties of the pollutants themselves, and the degradation efficiency varies with the change of molecular weight.

[0003] Existing studies have shown that bacteria-driven biodegradation is considered to be the most promising strategy among many remediation methods because of its complete degradation of pollutants, low treatment cost, high safety and low environmental disturbance. At present, a number of degradation strains with single PAEs degradation ability have been isolated and purified at the laboratory level. Some strains have shown good degradation effects in laboratory studies, but do not have broad-spectrum PAEs degradation capabilities. Soil is a natural microbial resource library, and single degrading bacteria lack competitiveness in highly complex indigenous bacterial communities. Therefore, it is extremely important to construct a multifunctional microbial community that is controllable, assemblable, has a clear interaction mechanism, stable functions and is competitive through scientific means in soil microbial remediation and improving plant production potential.

[0004] In recent years, although the idea of ​​synthetic microbial ecology has been established, there are not enough cases of synthetic microbial flora applied in actual production. As a resource product of agricultural waste, biochar has been used for soil improvement in recent years due to its superior physical and chemical properties and porous structure. The rich pore structure of biochar is considered to be an excellent microbial immobilization material, which not only provides a habitat for microorganisms, but also accelerates the degradation or adsorption of organic pollutants. Strains with disease resistance are selected to be compounded with PAEs degrading bacteria and loaded on biochar materials to form a competitive and stable synthetic flora.

[0005] Cucumber is a major vegetable in facility agriculture and occupies an important position in agricultural economy. It is usually planted in greenhouse facilities or covered with mulch. The different types and concentrations of PAEs residues in the soil environment cause stress on cucumber seedlings, growth and yield. In addition, PAEs have endocrine disrupting effects, posing a huge threat to human food health and safety. Therefore, the ecological safety of PAEs and the response of plants to their stress have attracted attention. Summary of the invention

[0006] The purpose of the present invention is to solve the problem that the current single degradation strain does not have the broad-spectrum degradation ability of PAEs, and the single degradation strain has an unstable PAEs degradation effect in the indigenous bacterial community, and to provide a method for preparing a biochar-loaded synthetic microbial agent and its application in plant growth promotion and soil remediation.

[0007] A method for preparing a biochar-loaded synthetic microbial agent is carried out according to the following steps:

[0008] Step S1: Pseudomonas sp. DNB-S1 culture;

[0009] Inoculate Pseudomonas sp. DNB-S1 into a LB medium containing a mixture of phthalates and dibutyl phthalate at an inoculation amount of 1 to 5%, and culture on a shaking platform at a temperature of 25 to 35° C. and a pH of 6 to 9 for 6 to 8 hours to obtain DNB-S1 bacteria;

[0010] Step S2: Cultivating Bacillus subtilis MES810;

[0011] Bacillus subtilis MES810 is inoculated into an LB medium containing a mixture of phthalates and dibutyl phthalate at an inoculation amount of 1 to 5%, and is cultured on a shaking platform at a temperature of 25 to 35° C. and a pH value of 6 to 9 for 6 to 8 hours to obtain MES810 bacteria;

[0012] Step S3: preparation of intermediate products;

[0013] The DNB-S1 cells in step S1 are cultured to OD=1, and the MES810 cells in step S2 are centrifuged, and then inoculated into a PBS buffer solution with a mass fraction of 0.5-1% biochar. After 6-8 hours of shaking, an intermediate product is obtained, and the volume ratio of DNB-S1 cells to the centrifuged MES810 cells is 1:1;

[0014] Step S4: preparation of biochar-loaded synthetic microbial agent;

[0015] The intermediate product in step S3 is inoculated into an inorganic salt culture medium containing a mixed substance composed of phthalates and dibutyl phthalate, with an inoculation amount of 1-5%, and cultured in a shaking table at a temperature of 25-35° C. and a pH value of 6-9 for 6-8 hours to obtain a biochar-loaded synthetic microbial agent.

[0016] The invention discloses an application of a biochar-loaded synthetic microbial agent, wherein the biochar-loaded synthetic microbial agent is used to degrade phthalates, dibutyl phthalate, dimethyl phthalate, diethyl phthalate or di(2-ethyl)hexyl phthalate in soil, and to promote cucumber growth and resist cucumber wilt.

[0017] Principle of the present invention:

[0018] The present invention selects microorganisms that are beneficial to plant growth as soil environmental conditioners, fully utilizes environmental pollutants as carbon sources and energy sources required for the growth of the microorganisms themselves, utilizes environmental pollutants and their secondary metabolites to carry out co-metabolism with other organic matter in the environment, and achieves functional and ecological stability of synthetic microbial communities in practical applications, guiding the practical application of synthetic bacterial communities from the two aspects of pollution control and plant growth promotion.

[0019] The present invention constructs a functionally stable and assemblable microbial community to achieve simultaneous restoration and production, and evaluates the ecological stability of the synthetic microbial community and the growth status of community members based on model and application verification; explores the growth-promoting effect of the multifunctional microbial community on cucumber; dynamically monitors the residual amount of soil PAEs and the content of PAEs in various parts of cucumber during the restoration process of the synthetic microbial community, evaluates the ability of the multifunctional synthetic bacterial community to block and control the absorption of soil PAEs by cucumber, and explains the key functions of the synthetic microbial community in driving soil health, plant health, and regulating the sustainable development of the "soil-stressed plant ecosystem" in practical applications.

[0020] The synthetic microbial agent constructed by the present invention exhibits more excellent performance in the degradation and utilization of DBP, and its 48-hour degradation rate of DBP is greater than the degradation rate when only DNB-S1 degradation bacteria are present. Among them, the pores of biochar provide a suitable environment for the habitation of microorganisms, which not only avoids the damage of high-concentration DBP to bacteria, but also promotes the utilization of DBP adsorbed on biochar by DNB-S1, which is beneficial to the chemotaxis of bacteria; the alkaline substances contained in biochar regulate the medium environment that gradually becomes acidic due to the increase of DBP secondary metabolites, and improve the stability of microbial growth and metabolism.

[0021] In the synthetic microbial agent constructed by the present invention, Bacillus subtilis MES810 enhances the efficient expression activity of the degradation gene of the DNB-S1 strain and increases the metabolic intensity of DNB-S1 to DBP. The efficient expression of the degradation gene of the DNB-S1 strain improves the response of the MES810 strain to the environment, creates a more favorable living space for the MES810 strain, and keeps the biocontrol characteristics of the MES810 strain stable under PAEs stress conditions. The DNB-S1 and MES810 strains selectively utilize strain metabolites such as simple organic carbon compounds to activate strain growth, strain degradation and the production of antibacterial substances.

[0022] Beneficial effects of the present invention:

[0023] The present invention provides a method for preparing a biochar-loaded synthetic microbial agent, which comprises a compound of DNB-S1 and MES810. In terms of growth characteristics, DNB-S1 can degrade 500 mg·L in 72 hours. -1 The biochar can degrade more than 95% of dibutyl phthalate (DBP); MES810 has a significant antibacterial effect on cucumber wilt. Laboratory tests show that the antibacterial rate of the cucumber wilt disease strain in the PDA solid medium is 55.7% after 7 days, and the inhibition diameter is 38.57mm. At the same time, the biochar load maintains the stability and efficient degradation characteristics of the synthetic microbial agent. After compounding, in the presence of 500mg·L -1 In the inorganic salt culture medium of DBP, the PAEs degradation capacity reaches more than 95% in 48 hours. It can be seen that the microbial agent synthesized by the present invention has a higher PAEs repair capacity and a stronger plant biocontrol capacity.

[0024] The multifunctional synthetic microbial agent prepared by the present invention can grow rapidly and stably in a culture medium containing PAEs, and is adaptable to a wide range of PAEs concentrations and a wide range of temperatures, is particularly suitable for the application of high-concentration PAEs, and has a high PAEs degradation efficiency.

[0025] The multifunctional synthetic microbial agent prepared by the present invention has a small overlap in ecological niches, and the community members (DNB-S1 and MES810) are in a stable symbiotic state during the synergistic degradation of PAEs.

[0026] The present invention can obtain a preparation method of a biochar-loaded synthetic microbial agent and the application of the biochar-loaded synthetic microbial agent in plant growth promotion and soil remediation. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1 A graph showing the degradation capabilities of the biochar-loaded synthetic microbial agent and DNB-S1 prepared in Example 1 on PAEs;

[0028] Figure 2A diagram showing the growth-promoting effect of the biochar-loaded synthetic microbial agent prepared in Example 1 on cucumber;

[0029] Figure 3 It shows the accumulation of PAEs in cucumbers under PAEs pollution conditions after the application of the biochar-loaded synthetic microbial agent prepared in Example 1;

[0030] Figure 4 A graph showing the residual PAEs in the soil after application of the biochar-loaded synthetic microbial agent prepared in Example 1 under PAEs contamination conditions. DETAILED DESCRIPTION

[0031] Specific implementation method 1: This implementation method is a method for preparing a biochar-loaded synthetic microbial agent, which is carried out according to the following steps:

[0032] Step S1: Pseudomonas sp. DNB-S1 culture;

[0033] Inoculate Pseudomonas sp. DNB-S1 into a LB medium containing a mixture of phthalates and dibutyl phthalate at an inoculation amount of 1 to 5%, and culture on a shaking platform at a temperature of 25 to 35° C. and a pH of 6 to 9 for 6 to 8 hours to obtain DNB-S1 bacteria;

[0034] Step S2: Cultivating Bacillus subtilis MES810;

[0035] Bacillus subtilis MES810 is inoculated into an LB medium containing a mixture of phthalates and dibutyl phthalate at an inoculation amount of 1 to 5%, and is cultured on a shaking platform at a temperature of 25 to 35° C. and a pH value of 6 to 9 for 6 to 8 hours to obtain MES810 bacteria;

[0036] Step S3: preparation of intermediate products;

[0037] The DNB-S1 cells in step S1 are cultured to OD=1, and the MES810 cells in step S2 are centrifuged, and then inoculated into a PBS buffer solution with a mass fraction of 0.5-1% biochar. After 6-8 hours of shaking, an intermediate product is obtained, and the volume ratio of DNB-S1 cells to the centrifuged MES810 cells is 1:1;

[0038] Step S4: preparation of biochar-loaded synthetic microbial agent;

[0039] The intermediate product in step S3 is inoculated into an inorganic salt culture medium containing a mixed substance composed of phthalates and dibutyl phthalate, with an inoculation amount of 1-5%, and cultured in a shaking table at a temperature of 25-35° C. and a pH value of 6-9 for 6-8 hours to obtain a biochar-loaded synthetic microbial agent.

[0040] Specific embodiment 2: The difference between this embodiment and specific embodiment 1 is that each 1L LB medium in step S1 and step S2 is composed of 10g·L -1 Tryptone, 5 g L -1 Yeast extract and 10 g L -1 Composition of sodium chloride.

[0041] The other steps are the same as those in the first specific implementation.

[0042] Specific embodiment 3: This embodiment differs from specific embodiment 1 or 2 in that the concentrations of phthalates and dibutyl phthalate in each 1L LB medium are both 100-500 mg·L -1 .

[0043] The other steps are the same as those in the first or second embodiment.

[0044] Specific embodiment 4: The difference between this embodiment and specific embodiments 1 to 3 is that the rotation speed of the shaking table in step S1 is 125 to 175 r / min. -1 .

[0045] The other steps are the same as those in Specific Embodiments 1 to 3.

[0046] Specific embodiment 5: The difference between this embodiment and specific embodiments 1 to 4 is that the rotation speed of the shaking table in step S2 is 125 to 175 r / min. -1 .

[0047] The other steps are the same as those in Specific Embodiments 1 to 4.

[0048] Specific embodiment 6: The difference between this embodiment and specific embodiments 1 to 5 is that the rotation speed of the shaking table in step S3 is 125 to 175 r / min. -1 .

[0049] The other steps are the same as those in Specific Embodiments 1 to 5.

[0050] Specific embodiment seven: The difference between this embodiment and any one of specific embodiments one to six is ​​that in step S4, each 1L of inorganic salt culture medium consists of 1g K2HPO4·3H2O, 0.4g MgSO4·7H2O, 1g NaCl, 0.075g CaCl2·2H2O, 0.01g FeCl3·6H2O and 0.5g NH4NO3, and the pH = 7.0.

[0051] The other steps are the same as those in Specific Embodiments 1 to 6.

[0052] Specific embodiment 8: This embodiment differs from specific embodiments 1 to 7 in that the concentrations of phthalates and dibutyl phthalate in each 1L of inorganic salt culture medium are both 100-500 mg·L -1 .

[0053] The other steps are the same as those in Specific Embodiments 1 to 7.

[0054] Specific embodiment 9: The difference between this embodiment and specific embodiments 1 to 8 is that the rotation speed of the shaking table in step S4 is 125 to 175 r / min. -1 .

[0055] The other steps are the same as those in Specific Embodiments 1 to 8.

[0056] Specific embodiment ten: This embodiment is an application of a biochar-loaded synthetic microbial agent, and the biochar-loaded synthetic microbial agent is used to degrade phthalates, dibutyl phthalate, dimethyl phthalate, diethyl phthalate or di(2-ethyl)hexyl phthalate in the soil, and to promote cucumber growth and resist cucumber wilt.

[0057] The following examples are used to verify the beneficial effects of the present invention:

[0058] Example 1: A method for preparing a biochar-loaded synthetic microbial agent, comprising the following steps:

[0059] Step S1: Pseudomonas sp. DNB-S1 culture;

[0060] Pseudomonas sp. DNB-S1 was inoculated into LB medium containing a mixture of phthalates and dibutyl phthalate at an inoculum of 3% and was cultured at 35°C and pH 7 at a rate of 125 r·min. -1 The cells were cultured on a shaker at a speed of 800 nm for 8 h to obtain DNB-S1 bacteria.

[0061] The Pseudomonas sp. DNB-S1 is deposited in the General Microbiology Center of China National Microbiological Culture Collection Administration, the deposit address is No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, the deposit date is November 6, 2014, and the deposit number is CGMCC No.9526.

[0062] Step S2: Cultivating Bacillus subtilis MES810;

[0063] Bacillus subtilis MES810 was inoculated into LB medium containing a mixture of phthalates and dibutyl phthalate at an inoculum size of 3%, and the mixture was grown at 125 r·min at a temperature of 30°C and a pH of 7. -1 The cells were cultured on a shaking incubator at a speed of 800 nm for 8 h to obtain MES810 bacteria.

[0064] The Bacillus subtilis MES810 is deposited in the General Microbiology Center (CGMCC) of the China Microorganism Culture Collection Administration, the deposit address is No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, the deposit date is August 10, 2017, and the deposit number is CGMCC No.14514.

[0065] In step S1 and step S2, each 1L of LB medium is composed of 10 g·L -1 Tryptone, 5 g L -1 Yeast extract and 10 g L -1 The concentration of phthalates and dibutyl phthalate in each 1L LB medium was 500 mg·L -1 ;

[0066] Step S3: preparation of intermediate products;

[0067] The DNB-S1 cells in step S1 were cultured to OD = 1, and the MES810 cells in step S2 were centrifuged and then inoculated into PBS buffer with a mass fraction of 1% biochar. -1 After 6 hours of shaking on a speed shaker, an intermediate product was obtained, and the volume ratio of DNB-S1 bacteria to the centrifuged MES810 bacteria was 1:1;

[0068] Step S4: preparation of biochar-loaded synthetic microbial agent;

[0069] The intermediate product in step S3 is inoculated into an inorganic salt culture medium containing a mixture of phthalates and dibutyl phthalate, with an inoculation amount of 3%, and cultured in a shaking table at a temperature of 30° C. and a pH value of 7 for 8 hours to obtain a biochar-loaded synthetic microbial agent;

[0070] In step S4, each 1L of inorganic salt culture medium contains 1g K2HPO4·3H2O, 0.4g MgSO4·7H2O, 1g NaCl, 0.075g CaCl2·2H2O, 0.01g FeCl3·6H2O and 0.5g NH4NO3, and the pH is 7.0; the concentrations of phthalates and dibutyl phthalate in each 1L of inorganic salt culture medium are both 500mg·L -1 .

[0071] 1. Determination of the synergistic degradation ability of synthetic microbial agents on PAEs:

[0072] Take the same volume of n-hexane as the extractant and mix it with the biochar-loaded synthetic microbial agent prepared in step S4, vortex and oscillate for 6 to 7 minutes, let it stand for stratification, take the upper organic solution, filter it with a funnel containing anhydrous sodium sulfate, pass the obtained filtrate through a 0.22μm Jinteng organic filter membrane, put the filtrate into a 1.5mL brown injection bottle, and determine the DBP concentration by gas chromatography-mass spectrometry (GC-MS). The results showed that the synthetic microbial agent maintained stability and efficient degradation characteristics. Figure 1 As shown, after compounding, in the presence of 500mg·L -1 In the inorganic salt culture medium of DBP, the PAEs degradation capacity reached more than 90% in 48 hours.

[0073] 2. Determination of the growth-promoting effect of synthetic microbial agents on cucumber:

[0074] Four treatment groups were selected: CK (control group), R1 (PAEs contaminated soil (20 mg kg -1 )、R2(PAEs contaminated soil (20 mg kg -1 )+Synthetic microbial agent (1×10 8 CFU·mL -1 )) and R3 (synthetic microbial agent (1×10 8 CFU·mL -1 )) The cucumbers planted were sampled at 35 days to measure the length (cm) and fresh weight (g) of the above-ground and underground parts of the plants. The roots were cleaned with distilled water and scanned with a root scanner. After the above indicators were measured, they were dried in an oven at 80°C to constant weight and then the dry weight (g) of the above-ground and underground parts was measured. Figure 2As shown in the results, the synthetic microbial agent promoted the growth of cucumber plants in diseased soil and reduced the incidence of cucumber wilt; it showed a positive effect on the growth and development of the root system, optimized the root structure, improved the health of the root system, and achieved the prevention and control effect of soil-borne diseases of cucumber.

[0075] 3. Accumulation of PAEs in cucumbers and residues of PAEs in soil:

[0076] Extraction method of PAEs in cucumber: The above-ground and underground parts of cucumber samples with different treatments were taken every 7 days. After freeze-drying, 0.2 g was accurately weighed and placed in a 10 mL glass tube. 3 mL of distilled water was added. After vortexing until uniform, an equal volume of n-hexane was added for extraction. After vortexing for 3 min, ultrasonic extraction was performed for 40 min. After removal, the sample was centrifuged (3000 r / min) -1 , 20min), the supernatant was filtered through a 0.22μm Jinteng organic filter membrane, placed in a 1.5mL brown injection bottle, and measured by GC-MS. Figure 3 As shown in Figure 2, at 35 days, the DBP residue in cucumber plants was less than 0.02 mg kg -1 .

[0077] Extraction method of PAEs in soil: 5 g soil sample was accurately weighed every 7 days, and dichloromethane was used as the extractant. After vortexing and ultrasonication, centrifugation (6000 r / min) was performed. -1 , 10min), collect the supernatant, filter it, and then concentrate it by rotary evaporation. Adjust the volume of the solution to 2mL and filter it through a 0.22μm Jinteng organic filter membrane. Put it into a 1.5mL brown injection bottle and measure it by GC-MS. Figure 4 As shown, at day 35, the DBP residue in the soil was less than 30%.

[0078] 4. Effect of synthetic microbial agents on cucumber resistance to Fusarium wilt:

[0079] Four treatment groups were selected: T0 (control soil), T1 (inoculated with cucumber wilt and 20 mg kg -1 DBP-infected soil), T2 (inoculated with cucumber wilt and 20 mg kg -1 DBP diseased soil + MES810) and T3 (inoculated with cucumber wilt and 20 mg kg -1 The diseased soil was prepared as follows: DBP (20 mg kg -1 ) was evenly sprayed on the soil and placed away from light for 24 hours until the acetone dissipated (evaporated to no obvious odor); Fusarium oxysporum was activated to a spore suspension concentration of 2×10 9 cfu·mL -1After filtering the mycelium, put it into sterile soil at a concentration of 1×10 8 cfu·mL -1 The diseased soil was transplanted. Cucumber seedlings with the same growth were transplanted and the growth and disease of cucumbers were observed. The disease incidence of the treatment group inoculated with synthetic microbial agents under the stress of DEHP and pathogenic bacteria was significantly reduced, and the disease prevention effect was significantly improved.

[0080] Table 1 shows the effect of synthetic microbial agents on cucumber resistance to Fusarium wilt;

[0081] Table 1

[0082] Treatment Group T0 T1 T2 T3 Incidence (%) 29.87±2.42 72.33±6.81 43.33±2.79 26.57±0.97 Disease index 14.02±3.47 62.53±4.97 21.93±3.26 10.37±3.78 Disease prevention effect (%) — — 64.93±3.17 83.42±0.90

[0083] Note: T0 is the control group soil, T1 group was inoculated with cucumber wilt and 20 mg kg -1 The diseased soil of DBP, T2 group was inoculated with cucumber wilt and 20 mg·kg -1 DBP+MES810, T3 group was inoculated with cucumber wilt and 20 mg·kg -1 DBP+synthetic microbial agent, the experiment was repeated 3 times.

Claims

1. A method for preparing a biochar-loaded synthetic microbial agent, characterized in that The preparation method is carried out according to the following steps: Step S1: Pseudomonas sp. DNB-S1 culture; Inoculate Pseudomonas sp. DNB-S1 into a LB medium containing a mixture of phthalates and dibutyl phthalate at an inoculation amount of 1 to 5%, and culture on a shaking platform at a temperature of 25 to 35° C. and a pH of 6 to 9 for 6 to 8 hours to obtain DNB-S1 bacteria; Step S2: Cultivating Bacillus subtilis MES810; Bacillus subtilis MES810 is inoculated into an LB medium containing a mixture of phthalates and dibutyl phthalate at an inoculation amount of 1 to 5%, and is cultured on a shaking platform at a temperature of 25 to 35° C. and a pH value of 6 to 9 for 6 to 8 hours to obtain MES810 bacteria; Step S3: preparation of intermediate products; The DNB-S1 cells in step S1 are cultured to OD=1, and the MES810 cells in step S2 are centrifuged, and then inoculated into a PBS buffer solution with a mass fraction of 0.5-1% biochar. After 6-8 hours of shaking, an intermediate product is obtained, and the volume ratio of DNB-S1 cells to the centrifuged MES810 cells is 1:1; Step S4: preparation of biochar-loaded synthetic microbial agent; The intermediate product in step S3 is inoculated into an inorganic salt culture medium containing a mixed substance composed of phthalates and dibutyl phthalate, with an inoculation amount of 1-5%, and cultured in a shaking table at a temperature of 25-35° C. and a pH value of 6-9 for 6-8 hours to obtain a biochar-loaded synthetic microbial agent.

2. The method for preparing a biochar-loaded synthetic microbial agent according to claim 1, characterized in that In step S1 and step S2, each 1L of LB medium is composed of 10 g·L -1 Tryptone, 5 g L -1 Yeast extract and 10 g L -1 Composition of sodium chloride.

3. The method for preparing a biochar-loaded synthetic microbial agent according to claim 1 or 2, characterized in that The concentration of phthalates and dibutyl phthalate in 1L LB medium is 100-500 mg / L -1 .

4. The method for preparing a biochar-loaded synthetic microbial agent according to claim 1, characterized in that The speed of the shaking table in step S1 is 125-175 r / min -1 .

5. The method for preparing a biochar-loaded synthetic microbial agent according to claim 1, characterized in that The speed of the shaking table in step S2 is 125-175 r / min -1 .

6. The method for preparing a biochar-loaded synthetic microbial agent according to claim 1, characterized in that The speed of the shaking table in step S3 is 125-175 r / min -1 .

7. The method for preparing a biochar-loaded synthetic microbial agent according to claim 1, characterized in that In step S4, each 1L of inorganic salt culture medium consists of 1g K2HPO4·3H2O, 0.4g MgSO4·7H2O, 1g NaCl, 0.075g CaCl2·2H2O, 0.01g FeCl3·6H2O and 0.5g NH4NO3, and the pH is 7.

0.

8. The method for preparing a biochar-loaded synthetic microbial agent according to claim 1 or 7, characterized in that The concentration of phthalates and dibutyl phthalate in each liter of inorganic salt culture medium is 100-500 mg·L -1 .

9. The method for preparing a biochar-loaded synthetic microbial agent according to claim 1, characterized in that The rotation speed of the shaking table in step S4 is 125 to 175 r / min. -1 .

10. Use of a biochar-loaded synthetic microbial agent prepared by the method according to any one of claims 1 to 9, characterized in that The biochar-loaded synthetic microbial agent is used in degrading phthalates, dibutyl phthalate, dimethyl phthalate, diethyl phthalate or di(2-ethyl)hexyl phthalate in soil, and promoting cucumber growth and resisting cucumber wilt.

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