A method for repairing cadmium contaminated soil by using arbuscular mycorrhizal fungi in combination with sweet sorghum

Abstract

The application discloses a method for repairing cadmium contaminated soil by using arbuscular mycorrhizal fungi and sweet sorghum, and belongs to the technical field of cadmium contaminated soil repair, and comprises the steps of arbuscular mycorrhizal fungi collection and induced culture, arbuscular mycorrhizal fungi single spore culture and cadmium tolerant strain screening, soil pretreatment and soil repair. After the soil is treated, the arbuscular mycorrhizal fungi and the sweet sorghum are combined to repair the cadmium contaminated soil, so that the problem of low infection rate of the arbuscular mycorrhizal fungi in the cadmium contaminated soil and low repair efficiency is solved.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of cadmium contaminated soil remediation, and particularly relates to a method for remediation of cadmium contaminated soil by using arbuscular mycorrhizal fungi in combination with sweet sorghum. BACKGROUND

[0002] In recent years, with the deepening of agricultural modernization, agriculture has made great achievements, and both quality and yield have been improved. However, while great development has been achieved, the soil and environment have been damaged to some extent, especially the heavy metal pollution of the soil is more serious, which reduces the fertility of the soil. The heavy metal pollution of the soil has become an important management project.

[0003] Cadmium is one of the most concerned elements in heavy metal pollution, and its compounds are all toxic substances. A very small amount of cadmium can cause harm to the human body. Once the cadmium in the soil is excessive, it is difficult to be removed, and after being absorbed by plants, it can damage the structure and function of plant chloroplasts, thereby inhibiting plant photosynthesis, damaging the antioxidant system, and leading to slow growth, yield and quality reduction of crops. At present, there are three types of cadmium contaminated soil remediation technologies in China. One is engineering remediation. Engineering remediation is complete and effective, but the engineering quantity is large, the investment is high, and the land structure itself is damaged. The second is physical and chemical remediation. The physical and chemical remediation method is also the mainstream method at present. Compared with engineering remediation, many physical and chemical technologies are in-situ remediation, which can effectively reduce the transportation cost. However, this method has high technical requirements, and the contaminated soil is difficult to be treated on a large scale. The third is biological remediation. The biological remediation method is a new technical means and is also considered as an effective means for future development. It mainly uses plants and microorganisms to reduce pollutants in the soil and purify or reduce the toxicity of heavy metals.

[0004] At present, many plants that can be used for cadmium contaminated soil remediation have been found, including Viola philippica, honeysuckle, and Solanum nigrum. However, the biomass of these plants is small, the remediation efficiency is low, and the remediation time is long, which is difficult to meet the production needs. Sweet sorghum is an annual plant of the Poaceae family, which has large biomass, strong stress resistance, and strong tolerance to heavy metals, and can well absorb cadmium ions in the soil, so it is an ideal plant for cadmium contaminated soil remediation.

[0005] Arbuscular mycorrhizal fungi are a kind of beneficial microorganisms widely distributed in soil ecosystems, which can form a symbiotic system with sweet sorghum, reduce the toxicity of cadmium to sweet sorghum, increase the absorption of cadmium in the aboveground part of sweet sorghum and the accumulation of cadmium in the root system of sweet sorghum, and thereby improve the remediation efficiency of sweet sorghum for cadmium contaminated soil. However, in cadmium contaminated soil, the infection intensity of ordinary arbuscular mycorrhizal fungi to sweet sorghum is low, which greatly reduces the remediation efficiency of cadmium contaminated soil. SUMMARY

[0006] Based on the above technical problems, the purpose of the present application is to provide a method for repairing cadmium contaminated soil by using arbuscular mycorrhizal fungi combined with sweet sorghum, which solves the problem of low infection intensity of arbuscular mycorrhizal fungi on sweet sorghum in cadmium contaminated soil, resulting in low soil repair efficiency.

[0007] The present application solves the above technical problems by the following technical means:

[0008] A method for repairing cadmium contaminated soil by using arbuscular mycorrhizal fungi combined with sweet sorghum, the method is as follows:

[0009] (1) Arbuscular mycorrhizal fungi collection and induced culture: when arbuscular mycorrhizal fungal spores mature, plant roots and rhizosphere soil with good growth vigor are dug in cadmium contaminated soil, the roots are cut into 1cm segments and mixed with the rhizosphere soil to obtain a soil sample; the substrate is mixed evenly with the soil sample to obtain an induced culture medium, then sweet sorghum seeds are sown in the induced culture medium, and then placed in a light culture room for culture for 3 months, and the spores of arbuscular mycorrhizal fungi on the roots of sweet sorghum plants are separated;

[0010] (2) Single spore culture of arbuscular mycorrhizal fungi and cadmium tolerant strain screening: the induced culture separated spores are subjected to single spore culture to obtain single spore cultured spores, which are washed and then inoculated into cadmium contaminated soil after picking up healthy spores, and after dark culture, the strain with the highest germination rate is selected, which is the cadmium tolerant arbuscular mycorrhizal fungal strain, ready for use;

[0011] (3) Soil pretreatment: the pre-repair soil is plowed, then watered to wet the soil surface, and then the accelerator is sprinkled and left to stand for 1 month to obtain the pre-repair soil;

[0012] (4) Soil repair: the cadmium tolerant arbuscular mycorrhizal fungal strain selected is propagated to obtain a cadmium tolerant arbuscular mycorrhizal fungal mixture and sweet sorghum seeds to be sown, and then sown together in the pre-repair soil for soil repair, and the growth of sweet sorghum plants is normally maintained.

[0013] Further, the arbuscular mycorrhizal fungi collection operation in step (1) is as follows:

[0014] When arbuscular mycorrhizal fungal spores mature, plants with good growth vigor are selected in cadmium contaminated soil, the dead branches and leaves around the plant roots and stones are removed, then the plant roots and rhizosphere soil are dug with a sterilized small shovel, the aboveground stems and leaves are cut off, the roots are retained, the roots are cut into 1cm segments, and the roots and rhizosphere soil are mixed evenly to obtain a soil sample.

[0015] Further, the specific operation of single spore culture in step (2) is as follows:

[0016] A: The spores separated after induction culture are extracted and placed in a petri dish containing distilled water, and stored at 4°C for 48-60 hours. Then, the inactivated and damaged spores are removed, and the clean, fresh, bright, full and robust spores are reserved for use;

[0017] B: The substrate is filled in the hole seedling tray, and water is sprayed to fully wet the surface of the substrate. The robust spores in step A are inoculated in the substrate, and then sweet sorghum seeds are sowed on the substrate. The culture is obtained after 40-50 days of illumination culture;

[0018] C: The culture is longitudinally cut into 1 / 4 of the whole for wet screening, and the amount of mycelium and spore propagation are checked. The culture without infection and spores is discarded, and the culture containing spores is reserved;

[0019] D: The substrate is added in the pot, the culture containing spores is moved into the pot, sweet sorghum seeds are sowed again, and then the culture is continued to be illuminated for 3 months. The spores are separated to obtain the spores after single spore culture.

[0020] Further, the dark culture temperature in step (2) is 26°C, and the culture time is 20 days.

[0021] Further, the specific operation of the propagation in step (4) is as follows:

[0022] The screened cadmium-tolerant arbuscular mycorrhizal fungus strain is inoculated in the pot containing the substrate, and then sweet sorghum seeds are sowed. The culture is illuminated in the illumination room for 100-120 days. After the culture is completed, the sweet sorghum seeds are collected to obtain the seeds to be sowed. Then, the sweet sorghum root system in the pot is taken out, crushed and returned to the pot to mix with the substrate, to obtain the cadmium-tolerant arbuscular mycorrhizal fungus mixture.

[0023] Further, the promoter in step (3) includes the following raw materials by weight:

[0024] 3-5 parts of barium sulfate, 30-50 parts of dimethyl formamide, 80-120 parts of magnesium stearate, 80-120 parts of maltose, 80-120 parts of L-methionine, and 600-900 parts of wheat germ peptide.

[0025] Further, the preparation method of the promoter is as follows:

[0026] S1: The barium sulfate and dimethyl formamide are mixed, stirred at a speed of 500-800 r / min for 5-10 min to obtain a barium sulfate mixed solution for standby;

[0027] S2: 60wt% ethanol solution is added to the magnesium stearate and mixed uniformly. Then, the magnesium stearate solution is obtained by heating in hot water for standby;

[0028] S3: maltose, L-methionine, wheat germ peptide are mixed, then water is added to stir and mix uniformly, then barium sulfate mixed solution and magnesium stearate solution are added, and the mixture is homogenized in an ultrasonic homogenizer to obtain the accelerator.

[0029] Further, the temperature of the hot water in the water bath heating in step S2 is 60-90 DEG C, and the water bath heating time is 8-15 min.

[0030] Further, the homogenization conditions in step S3 are: frequency 20-25 kHz, temperature 25-35 DEG C, and homogenization time 5-15 min.

[0031] Further, the application amount of the accelerator is 30-100 g / m 2 .

[0032] Further, the medium used in steps (1) arbuscular mycorrhizal fungi collection and induced culture, (2) arbuscular mycorrhizal fungi single spore culture and cadmium-resistant strain screening, and (4) soil remediation is a mixture of river sand and zeolite in a weight ratio of 1:1, which is sterilized at high temperature to obtain.

[0033] Further, the light culture conditions in steps (1) arbuscular mycorrhizal fungi collection and induced culture, (2) arbuscular mycorrhizal fungi single spore culture and cadmium-resistant strain screening, and (4) soil remediation are: light intensity 2000-25000 lx, and temperature 22-30 DEG C.

[0034] In cadmium-contaminated soil, arbuscular mycorrhizal fungi can promote sweet sorghum to absorb cadmium elements, but high concentration of cadmium can cause arbuscular mycorrhizal fungi to be stressed by cadmium, thereby reducing the infection rate of sweet sorghum, so that ordinary planting process is difficult to use sweet sorghum to repair cadmium-contaminated land, thereby reducing the soil remediation efficiency, therefore, it is necessary to improve the infection rate of arbuscular mycorrhizal fungi to sweet sorghum in cadmium-contaminated soil, thereby improving the remediation efficiency of sweet sorghum to cadmium-contaminated soil.

[0035] The present application collects arbuscular mycorrhizal fungi, then performs induced culture, single spore culture, screening and propagation to obtain a cadmium-resistant arbuscular mycorrhizal fungi mixture and sweet sorghum seeds to be planted, thereby improving the cadmium resistance of arbuscular mycorrhizal fungi and the affinity of arbuscular mycorrhizal fungi to sweet sorghum, and further improving the infection rate of arbuscular mycorrhizal fungi to sweet sorghum in cadmium-contaminated soil and the remediation efficiency.

[0036] The magnesium stearate, maltose and L-methionine in the accelerator are entered into the soil to increase the soil permeability and improve the soil physical and chemical properties, so that the arbuscular mycorrhizal fungi can grow and reproduce in the soil, the number is rapidly increased, and the infection rate of sweet sorghum is increased. However, the magnesium stearate, maltose and L-methionine in the soil are easily decomposed, and the action time is short, therefore, the wheat germ peptide is added to the accelerator to increase the stability of the magnesium stearate, maltose and L-methionine in the soil and prolong the action time. At the same time, the wheat germ peptide can also be used as a nutrient material for earthworms in the soil to promote the growth and reproduction of earthworms, increase the number of earthworms and the life activities of earthworms in the soil, further improve the soil permeability and improve the soil physical and chemical properties, and further improve the arbuscular mycorrhizal fungal infection rate.

[0037] After the arbuscular mycorrhizal fungi infect the sweet sorghum, the sweet sorghum needs to continuously provide carbon source and other substances for the arbuscular mycorrhizal fungi, but the photosynthesis of the sweet sorghum is inhibited in the cadmium contaminated soil, so that the carbon source content in the plant is reduced, and the growth and reproduction of the arbuscular mycorrhizal fungi are affected. Therefore, barium sulfate is also added to the accelerator, the barium sulfate can promote the dissolution of organic phosphorus by phosphorus-dissolving microorganisms in the soil, increase the absorption of organic phosphorus by the sweet sorghum, strengthen the photosynthesis of the sweet sorghum, and then increase the accumulation amount of carbon source in the sweet sorghum, to meet the growth and reproduction needs of the arbuscular mycorrhizal fungi. With the continuous growth of the sweet sorghum, the root system is increasingly large, the amount of arbuscular mycorrhizal fungal infection is continuously increased, and the ability of the sweet sorghum to absorb cadmium ions is continuously improved, thereby improving the repair efficiency of the cadmium contaminated soil.

[0038] Advantages:

[0039] 1. The arbuscular mycorrhizal fungi are collected, induced and cultured, and then screened to obtain cadmium-resistant arbuscular mycorrhizal fungal strains, which are combined with sweet sorghum to repair the cadmium contaminated soil after propagation, so that the cadmium contaminated soil can be effectively repaired and the soil properties can be improved.

[0040] 2. Before repairing the cadmium contaminated soil, the soil is treated by adding an accelerator to the soil, and then the arbuscular mycorrhizal fungi are combined with sweet sorghum to repair the cadmium contaminated soil, so that the arbuscular mycorrhizal fungal infection rate can be effectively improved, and the repair efficiency of the cadmium contaminated soil can be improved. BRIEF DESCRIPTION OF DRAWINGS

[0041] Figure 1 : It is a mixture of the cadmium-resistant arbuscular mycorrhizal fungi in the experimental group 1 of the application after propagation;

[0042] Figure 2 : It is a diagram of the infection of arbuscular mycorrhizal fungi on sweet sorghum in the experimental group 1 of the application. DETAILED DESCRIPTION

[0043] The application will be described in detail below in conjunction with specific embodiments and drawings:

[0044] Example 1: Preparation of the accelerator

[0045] Take 3g of barium sulfate, 30g of dimethylformamide, 80g of magnesium stearate, 80g of maltose, 80g of L-methionine, and 600g of wheat germ peptide.

[0046] Preparation method:

[0047] S1: After mixing barium sulfate and dimethylformamide, stir at a speed of 500 r / min for 5 min to obtain a barium sulfate mixture for standby;

[0048] S2: Add 800g of 60wt% ethanol solution to the magnesium stearate and mix evenly, then place in 60℃ hot water for water bath heating for 8 min to obtain a magnesium stearate solution for standby;

[0049] S3: Mix maltose, L-methionine, and wheat germ peptide, then add 7.6kg of water and stir to mix evenly, then add the barium sulfate mixture and the magnesium stearate solution, and put into an ultrasonic homogenizer, homogenize at a frequency of 20kHz and a temperature of 25℃ for 5 min to obtain the accelerator.

[0050] Example 2: Preparation of the accelerator

[0051] Take 4g of barium sulfate, 40g of dimethylformamide, 100g of magnesium stearate, 100g of maltose, 100g of L-methionine, and 750g of wheat germ peptide.

[0052] Preparation method:

[0053] S1: After mixing barium sulfate and dimethylformamide, stir at a speed of 600 r / min for 8 min to obtain a barium sulfate mixture for standby;

[0054] S2: Add 1kg of 60wt% ethanol solution to the magnesium stearate and mix evenly, then place in 75℃ hot water for water bath heating for 12 min to obtain a magnesium stearate solution for standby;

[0055] S3: Mix maltose, L-methionine, and wheat germ peptide, then add 9.5kg of water and stir to mix evenly, then add the barium sulfate mixture and the magnesium stearate solution, and put into an ultrasonic homogenizer, homogenize at a frequency of 22kHz and a temperature of 30℃ for 10 min to obtain the accelerator.

[0056] Example 3: Preparation of the accelerator

[0057] Take 5g of barium sulfate, 50g of dimethylformamide, 120g of magnesium stearate, 120g of maltose, 120g of L-methionine, and 900g of wheat germ peptide.

[0058] Preparation method:

[0059] S1: After mixing barium sulfate with dimethylamide, stirring at 800 r / min for 10 min, a barium sulfate mixture was obtained and used as needed;

[0060] S2: 1.2 kg of 60 wt% ethanol solution was added to the magnesium stearate and mixed evenly, then placed in 90°C hot water for water bath heating for 12 min to obtain a magnesium stearate solution for standby;

[0061] S3: After mixing maltose, L-methionine and wheat germ peptide, 11.4 kg of water was added and stirred and mixed evenly, then the barium sulfate mixture and the magnesium stearate solution were added, and placed in an ultrasonic homogenizer under the conditions of a frequency of 25 kHz and a temperature of 35°C for 15 min to obtain the accelerator.

[0062] Comparative Example 1: Preparation of accelerator

[0063] Comparative to Example 1, the only difference is that no barium sulfate mixture is added during the preparation of the accelerator.

[0064] Preparation method:

[0065] 80 g of magnesium stearate, 80 g of maltose, 80 g of L-methionine and 600 g of wheat germ peptide were weighed.

[0066] S1: 800 g of 60 wt% ethanol solution was added to the magnesium stearate and mixed evenly, then placed in 60°C hot water for water bath heating for 8 min to obtain a magnesium stearate solution for standby;

[0067] S2: After mixing maltose, L-methionine and wheat germ peptide, 7.6 kg of water was added and stirred and mixed evenly, then the magnesium stearate solution was added, and placed in an ultrasonic homogenizer under the conditions of a frequency of 20 kHz and a temperature of 25°C for 5 min to obtain the accelerator.

[0068] Comparative Example 2: Preparation of accelerator

[0069] Comparative to Example 1, the only difference is that no magnesium stearate mixture, maltose and L-methionine are added during the preparation of the accelerator.

[0070] Preparation method:

[0071] 3 g of barium sulfate, 30 g of dimethylformamide and 600 g of wheat germ peptide were weighed.

[0072] S1: After mixing barium sulfate with dimethylamide, stirring at 500 r / min for 5 min, a barium sulfate mixture was obtained and used as needed;

[0073] S2: Wheat germ peptide was added to 6 kg of water and stirred to mix evenly, then the barium sulfate mixed solution and magnesium stearate solution were added, and the mixture was placed in an ultrasonic homogenizer at a frequency of 20 kHz and a temperature of 25°C for 5 min to obtain the accelerator.

[0074] Comparative Example 3: Preparation of an accelerator

[0075] Comparative Example 1 was formed, the only difference being that no wheat germ peptide was added during preparation of the accelerator.

[0076] Preparation method:

[0077] 3 g of barium sulfate, 30 g of dimethylformamide, 80 g of magnesium stearate, 80 g of maltose, and 80 g of L-methionine were weighed.

[0078] Preparation method:

[0079] S1: After mixing barium sulfate and dimethylformamide, stirring at a speed of 500 r / min for 5 min, a barium sulfate mixed solution was obtained and reserved for use;

[0080] S2: 800 g of 60 wt% ethanol solution was added to the magnesium stearate and mixed evenly, then placed in hot water at 60°C for 8 min to obtain a magnesium stearate solution, which was reserved for use;

[0081] S3: Maltose and L-methionine were mixed and then added to 1.6 kg of water and stirred to mix evenly, then the barium sulfate mixed solution and magnesium stearate solution were added, and the mixture was placed in an ultrasonic homogenizer at a frequency of 20 kHz and a temperature of 25°C for 5 min to obtain the accelerator.

[0082] After the preparation of the accelerator, the matrix was further prepared as follows:

[0083] River sand and zeolite were mixed in a weight ratio of 1:1, then water was added to wet them, and then they were placed in an autoclave and sterilized at 120°C for 1 h, with three continuous sterilizations, each with an interval of 24 hours, to obtain the matrix.

[0084] Example 4: Method for repairing cadmium-contaminated soil by arbuscular mycorrhizal fungi combined with sweet sorghum

[0085] (1) Arbuscular mycorrhizal fungi collection and induced culture: Select a cadmium contaminated soil in Baoji, Shaanxi as the collection site. When the spores of arbuscular mycorrhizal fungi mature, select spinach plants with good growth, remove the dead branches and leaves around the plant roots and stones, then use a sterilized small shovel to dig the spinach plant roots and rhizosphere soil, cut off the stems and leaves, leave the roots, cut the roots into small pieces of about 1 cm, mix the roots with the rhizosphere soil evenly to obtain the soil sample; Mix the substrate and the soil sample in a volume ratio of 1:1 to obtain the induced culture medium; Fill the induced culture medium into the flowerpot to 3 / 4 of the height, spray water to fully wet the surface of the medium, then select intact and undamaged sweet sorghum seeds with full particles for sowing, 10 seeds per pot, with a sowing depth of about 1.5 cm, then place it in a light culture room, cultivate for 3 months under the conditions of light intensity of 2000lx and temperature of 22℃, isolate the spores of arbuscular mycorrhizal fungi on the roots of sweet sorghum plants;

[0086] (2) Single spore culture of arbuscular mycorrhizal fungi and cadmium tolerant strain screening:

[0087] A: Extract the spores separated after induced culture using the Cobb wet sieve method, place them in a petri dish containing sterilized distilled water, store them at 4℃ for 48h, then wash and remove inactive and damaged spores, and reserve clean, fresh, shiny, full and healthy spores for use;

[0088] B: Fill the substrate into the hole seedling tray to 2 / 3 of the height of the hole seedling tray, spray water to fully wet the surface of the substrate, then inoculate the healthy spores in step A into the substrate, 1 spore per hole, cover 1 cm of substrate on the spores, then select intact and undamaged sweet sorghum seeds with full particles for sowing on the substrate, 3 seeds per hole, with a sowing depth of about 1.5 cm, then place it in a light culture room, cultivate for 45 days under the conditions of light intensity of 2000lx and temperature of 22℃ to obtain the culture;

[0089] C: Take out the culture and place it in a sterilized tray, use a sterilized wallpaper knife to cut 1 / 4 of the whole culture longitudinally for Cobb wet sieving, check the amount of mycelium and spore reproduction, discard the culture without infection and spores, and reserve the culture containing spores;

[0090] D: Fill the substrate into the flowerpot to 1 / 3 of the flowerpot, move the culture containing spores into the flowerpot, select the full and intact sweet sorghum seeds to reseed in the flowerpot, 5 per pot, the seeding depth is about 1.5 cm, then place in the light culture room, under the condition of light intensity of 2000 lx and temperature of 22℃, light culture for 3 months, then separate the spores to obtain the spores after single spore culture; dig the cadmium contaminated soil and fill it into the flowerpot, pick up the healthy spores after washing the spores after single spore culture and inoculate them into the flowerpot filled with cadmium contaminated soil, dark culture for 20 days at 26℃, then select the highest germination rate of the fungus, which is the cadmium tolerant arbuscular mycorrhizal fungus, for use;

[0091] (3) Soil pretreatment: plough the pre-repair soil, the ploughing thickness is about 25 cm, after ploughing, sprinkle water to fully soak the soil surface, then spray the accelerator prepared in example 1 into the pre-repair soil at the amount of 50 g / m 2 , stand for 1 month to obtain the treated pre-repair soil;

[0092] (4) Soil repair: fill the substrate into the flowerpot to 2 / 3, evenly lay the cadmium tolerant arbuscular mycorrhizal fungus selected on the substrate, cover with about 2 cm thick substrate, water the substrate to fully soak the surface, then select the full and intact sweet sorghum seeds for seeding, 5 per pot, the seeding depth is about 1.5 cm, then place in the light culture room, culture for 110 days under the condition of light intensity of 2000 lx and temperature of 22℃; after the culture is completed, collect the sweet sorghum seeds to obtain the seeds to be sowed, then cut off the stem and leaf parts and reserve the roots, crush the roots and put them back into the pot to mix with the substrate to obtain the cadmium tolerant arbuscular mycorrhizal fungus mixture; mix the cadmium tolerant arbuscular mycorrhizal fungus mixture and the seeds to be sowed according to the weight ratio of 20:1, then evenly sprinkle them into the treated pre-repair soil for soil repair, and normally maintain the growth of sweet sorghum plants.

[0093] Example 4: Arbuscular mycorrhizal fungus combined with sweet sorghum for repairing cadmium contaminated soil

[0094] In comparison with example 4, the only difference is that in step (2), the arbuscular mycorrhizal fungus is not put into the cadmium contaminated soil for screening, but is put into the ordinary soil for screening to obtain the arbuscular mycorrhizal fungus, and the rest of the steps are the same as example 4, and step (2) is as follows:

[0095] Single spore culture of arbuscular mycorrhizal fungus and screening of cadmium tolerant fungus

[0096] A: extract the separated spores after induction culture by using kob wet sieve method, place them in a petri dish containing sterilized distilled water, store at 4℃ for 48 h, then wash and remove the inactivated and damaged spores, reserve the clean, fresh, bright, full and healthy spores for use;

[0097] B: Fill the substrate into the plug tray to 2 / 3 of the plug tray, spray water to make the surface of the substrate fully wet, then inoculate the healthy spores in step A into the substrate, put 1 spore in each hole, cover 1 cm substrate on the spores, then screen the full and intact sweet sorghum seeds and sow them on the substrate, 3 seeds per hole, the sowing depth is about 1.5 cm, then place it in the light culture room, cultivate for 45 days under the condition of light intensity of 2000 lx and temperature of 22℃ to obtain the culture;

[0098] C: Take out the culture and place it in the sterilized tray, longitudinally cut 1 / 4 of the whole culture with a sterilized wallpaper knife to perform the Cobb wet screening, check the mycelium amount and spore reproduction, discard the culture without infection and spores, and reserve the culture containing spores;

[0099] D: Fill the substrate into the flowerpot to 1 / 3 of the flowerpot, move the culture containing spores into the flowerpot, screen the full and intact sweet sorghum seeds and sow them in the flowerpot again, 5 seeds per pot, the sowing depth is about 1.5 cm, then place it in the light culture room, cultivate for 3 months under the condition of light intensity of 2000 lx and temperature of 22℃, then separate the spores to obtain the spores after single spore culture; fill the ordinary soil into the flowerpot, wash the spores after single spore culture, pick up the healthy spores, inoculate them into the flowerpot filled with ordinary soil, cultivate in dark for 20 days at 26℃, then select the highest germination rate of the strains to obtain the arbuscular mycorrhizal fungus strains, for use;

[0100] Comparative Example 5: Method for repairing cadmium contaminated soil by arbuscular mycorrhizal fungi combined with sweet sorghum

[0101] In comparison with Example 4, the only difference is that the step (1) of the preparation of the arbuscular mycorrhizal fungi does not perform the induction culture, and the step (2) does not perform the single spore culture, but directly collects the arbuscular mycorrhizal fungi to screen the cadmium tolerant arbuscular mycorrhizal fungus strains, and the rest steps are the same as those of Example 4, and the steps (1) and (2) are as follows:

[0102] (1) Arbuscular mycorrhizal fungi collection: select a cadmium contaminated soil in a facility agriculture in Baoji, Shaanxi as the collection site, when the arbuscular mycorrhizal fungi spores mature, select the well-grown spinach plants, remove the dead branches and leaves and stones around the plant roots, then use the sterilized small shovel to dig the plant roots and rhizosphere soil, cut off the aboveground stems and leaves, leave the roots, cut the roots into small pieces of about 1 cm, mix the roots with the rhizosphere soil to obtain the soil sample;

[0103] (2) Cadmium tolerant strain screening: dig the cadmium contaminated soil, mix the soil sample with the cadmium contaminated soil according to the volume ratio of 1:1, fill it into the flowerpot, cultivate in dark for 20 days at 26℃, then select the highest germination rate of the strains to obtain the cadmium tolerant arbuscular mycorrhizal fungus strains, for use.

[0104] Example 6: Method for repairing cadmium contaminated soil by arbuscular mycorrhizal fungi combined with sweet sorghum

[0105] In comparison with Example 4, the difference lies in that in step (3), the soil is not treated with the promoting agent, but is directly treated after spraying clean water on the soil, and the rest of the steps are the same as those in Example 4. The soil pretreatment operation is specifically as follows:

[0106] (3) Soil pretreatment: the pre-repair soil is ploughed to a depth of about 25 cm, and then watered to fully soak the soil surface. Then, clean water is sprayed on the pre-repair soil in an amount of 50 g / m 2 , and the treated pre-repair soil is obtained after standing for 1 month;

[0107] Example 7: Method for repairing cadmium contaminated soil by arbuscular mycorrhizal fungi combined with sweet sorghum

[0108] In comparison with Example 4, the difference lies in that in step (1), the induction culture is not performed, and in step (2), the single spore culture is not performed, and the screening is not performed in the cadmium contaminated soil, but in the ordinary soil, and the soil is treated with clean water instead of the promoting agent. The specific steps are as follows:

[0109] (1) Arbuscular mycorrhizal fungi collection: select a cadmium contaminated soil in a facility agriculture in Baoji, Shaanxi as the collection site. When the arbuscular mycorrhizal fungal spores mature, select the spinach plants with good growth, remove the dead branches and leaves and stones around the plant roots, then use a sterilized small shovel to dig the plant roots and rhizosphere soil, cut off the aboveground stems and leaves, leave the roots, cut the roots into small pieces of about 1 cm, mix the roots with the rhizosphere soil evenly to obtain the soil sample;

[0110] (2) Strain screening: dig the ordinary soil, mix the soil sample with the ordinary soil according to a volume ratio of 1:1, then put it into a flowerpot, and cultivate it in dark at 26°C for 20 days. Then select the strain with the highest germination rate to obtain the arbuscular mycorrhizal fungal strain for later use;

[0111] (3) Soil pretreatment: the pre-repair soil is ploughed to a depth of about 25 cm, and then watered to fully soak the soil surface. Then, clean water is sprayed on the pre-repair soil in an amount of 50 g / m 2 , and the treated pre-repair soil is obtained after standing for 1 month;

[0112] (4) Soil remediation: fill the flowerpot with substrate to 2 / 3, evenly spread the screened arbuscular mycorrhizal fungi on the substrate, cover with about 2cm thick substrate, water to fully soak the surface of the substrate, then screen the sweet sorghum seeds with plump and intact particles for sowing, 5 per pot, the sowing depth is about 1.5cm, then place it in the light culture room, cultivate for 110 days under the condition of light intensity of 2000lx and temperature of 22℃; after the cultivation is completed, the sweet sorghum seeds are collected to obtain the to-be-sown sweet sorghum seeds, then the stem and leaf parts are cut off to retain the roots, the roots are crushed and then put back into the pot to mix with the substrate to obtain the arbuscular mycorrhizal fungi mixture; the arbuscular mycorrhizal fungi mixture and the to-be-sown sweet sorghum seeds are mixed in a weight ratio of 20:1, then evenly spread into the treated pre-remediation soil for soil remediation, and the growth of the sweet sorghum plant is normally maintained.

[0113] Experiment: Arbuscular mycorrhizal fungi combined with sweet sorghum for remediation of cadmium contaminated soil

[0114] 1. The promoters prepared in Example 1 and Comparative Examples 1-3 are subjected to the experiment of arbuscular mycorrhizal fungi combined with sweet sorghum for remediation of cadmium contaminated soil, and the experiment is divided into 8 groups: experimental group 1, control groups 1-6, and blank control group. The experiment is carried out in a cadmium-exceeding field in Baoji, Shaanxi. Before the experiment, the initial content of cadmium in the soil of the experimental site is measured to be 0.93mg / kg. The arbuscular mycorrhizal fungi collected from the cadmium contaminated soil are identified as Glomus mosseae.

[0115] 2. Promoter

[0116] Experimental group 1: the promoter prepared in Example 1 is selected;

[0117] Control groups 1-3: the promoters prepared in Comparative Examples 1-3 are selected respectively;

[0118] Control groups 4-5: the promoter prepared in Example 1 is selected;

[0119] Control group 6: water is selected;

[0120] Control group 7 (blank control): water is selected as the control.

[0121] 3. Method of arbuscular mycorrhizal fungi combined with sweet sorghum for remediation of cadmium contaminated soil

[0122] Experimental group 1: the method of arbuscular mycorrhizal fungi combined with sweet sorghum for remediation of cadmium contaminated soil in Example 4 is selected;

[0123] Control groups 1-3: the method of arbuscular mycorrhizal fungi combined with sweet sorghum for remediation of cadmium contaminated soil in Example 4 is selected;

[0124] Control groups 4-6: the methods of arbuscular mycorrhizal fungi combined with sweet sorghum for remediation of cadmium contaminated soil in Comparative Examples 4-6 are selected respectively;

[0125] Control group 7 (blank control group): The method for repairing cadmium contaminated soil by using the arbuscular mycorrhizal fungus combined with sweet sorghum of the comparative example 7 is selected as the control.

[0126] 4. Cadmium contaminated soil repair capacity detection: After the sweet sorghum is harvested, the cadmium ion concentration in each group of soil is detected again to obtain the soil cadmium content, and the arbuscular mycorrhizal fungus infection rate is determined by the trypan blue staining-microscope observation method, repeated 3 times, and the average data is shown in Table 1:

[0127] Table 1

[0128]

[0129] According to the data analysis of Table 1, it can be seen that:

[0130] (1) Compared with control groups 1-6 and the blank control group, the cadmium ion content in the soil repaired by experimental group 1 is lower than that of control groups 1-6 and the blank control group, which shows that after the arbuscular mycorrhizal fungus is collected, induced and cultured, single spore cultured and screened, and combined with sweet sorghum to repair the cadmium contaminated soil treated with the promoter, the repair efficiency can be effectively improved, and the cadmium ion content in the soil can be reduced.

[0131] (2) Compared with control groups 1-6 and the blank control group, the infection rate of experimental group 1 is significantly higher than that of control groups 1-6 and the blank control group, and compared with experimental group 1, the infection rate of control group 1 is reduced by 9.01%, which is due to the fact that the barium sulfate mixed solution is not added during the preparation of the promoter of control group 1, the accumulation of sweet sorghum carbon source is reduced, which has an adverse effect on the growth and reproduction of arbuscular mycorrhizal fungus, resulting in a decrease in the infection rate of arbuscular mycorrhizal fungus;

[0132] (3) Compared with experimental group 1, the infection rate of control group 2 is reduced by 12.79%, which is due to the fact that the magnesium stearate mixed solution, maltose and L-methionine are not added during the preparation of the promoter of control group 2, resulting in poor soil permeability and poor physicochemical properties, and the infection rate of arbuscular mycorrhizal fungus is reduced;

[0133] (4) Compared with experimental group 1, the infection rate of control group 3 is reduced by 11.90%, which is due to the fact that wheat germ peptide is not added during the preparation of the promoter of control group 3, resulting in a decrease in the effect of magnesium stearate, maltose and L-methionine, and a decrease in the number of earthworms and their life activities in the soil, resulting in poor soil physicochemical properties and a decrease in the infection rate of arbuscular mycorrhizal fungus.

[0134] (5) Compared with the experimental group 1, the infection rate of the control group 4 is reduced by 8.31%, which is due to the fact that the arbuscular mycorrhizal fungi in the control group 4 are not screened in the cadmium contaminated soil, but in the ordinary soil, resulting in that the adaptability of the arbuscular mycorrhizal fungi to cadmium is low, and the infection rate is reduced due to the toxic effect of cadmium ions when the arbuscular mycorrhizal fungi enter the cadmium contaminated soil;

[0135] (6) Compared with the experimental group 1, the infection rate of the control group 5 is reduced by 7.69%, which is due to the fact that the arbuscular mycorrhizal fungi in the control group 5 are not induced and cultured after being collected, and the affinity of the arbuscular mycorrhizal fungi to sweet sorghum is relatively low, resulting in that the infection rate is reduced;

[0136] (7) Compared with the experimental group 1, the infection rate of the control group 6 is reduced by 13.87%, which is due to the fact that the soil in the control group 6 is not treated with the promoter, but is directly repaired after being sprayed with water, and the soil permeability is insufficient, the carbon source accumulation of sweet sorghum is reduced, and the infection rate of the arbuscular mycorrhizal fungi is reduced, which shows that after the arbuscular mycorrhizal fungi are collected, induced and cultured, and screened, and then inoculated in the soil treated with the promoter and combined with sweet sorghum, the cadmium contaminated soil can be repaired, and the infection rate of the arbuscular mycorrhizal fungi can be effectively improved, and the problem of low infection rate of the arbuscular mycorrhizal fungi in the cadmium contaminated soil, resulting in low repair effect, can be solved.

[0137] The above examples are only used to illustrate the technical solutions of the present application, but not limit the present application. Although the present application has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that the technical solutions of the present application can be modified or replaced by equivalents without departing from the spirit and scope of the present application, and all of them should be covered in the scope of the claims of the present application. The technical, shape and structure parts not described in detail in the present application are all known technologies.

Claims

1. A method for remediating cadmium contaminated soil using arbuscular mycorrhizal fungi in combination with sweet sorghum, characterized in that, The method is as follows: (1) mycorrhizal fungi collection and induced culture: when the spores of mycorrhizal fungi mature, the well-growing plant roots and rhizosphere soil in cadmium-contaminated soil are collected, the roots are cut into 1 cm segments and mixed with the soil to obtain a soil sample; the substrate is mixed with the soil sample to obtain an induced culture medium, then sweet sorghum seeds are sown in the induced culture medium, and then placed in a light culture room for culture for 3 months, and the spores of mycorrhizal fungi on the roots of sweet sorghum plants are isolated; (2) single spore culture of mycorrhizal fungi and screening of cadmium-resistant strains: the spores isolated by induced culture are subjected to single spore culture to obtain spores after single spore culture, and then the healthy spores are picked up after washing and inoculated in cadmium-contaminated soil, and the strain with the highest germination rate is selected after dark culture, which is the cadmium-resistant mycorrhizal fungi strain, ready for use; (3) soil pretreatment: the pre-repair soil is plowed, then watered to wet the soil surface, and then the accelerator is sprayed to treat for 1 month to obtain the pre-repair soil; (4) soil remediation: the cadmium-resistant mycorrhizal fungi strain selected is propagated to obtain a cadmium-resistant mycorrhizal fungi mixture and sweet sorghum seeds to be sown, and then sown together in the pre-repair soil for soil remediation, and the growth of the sweet sorghum plants is normally maintained; The specific operation of propagation in step (4) is as follows: Inoculate the cadmium-resistant mycorrhizal fungi strain selected in the pot with substrate, then sow sweet sorghum seeds, and place them in a light culture room for light culture for 100-120 days; after the culture is completed, the sweet sorghum seeds are collected to obtain the sweet sorghum seeds to be sown; then the sweet sorghum roots in the pot are taken out and crushed and then put back into the pot and mixed with the substrate to obtain a cadmium-resistant mycorrhizal fungi mixture; The accelerator in step (3) includes the following raw materials by weight: 3-5 parts of barium sulfate, 30-50 parts of dimethylformamide, 80-120 parts of magnesium stearate, 80-120 parts of maltose, 80-120 parts of L-methionine, and 600-900 parts of wheat germ peptide.

2. The method for repairing cadmium contaminated soil by using arbuscular mycorrhizal fungi combined with sweet sorghum according to claim 1, characterized in that, The specific operation of single spore culture in step (2) is as follows: A: The spores isolated after induced culture are placed in a petri dish containing distilled water, stored at 4°C for 48-60h, then washed and the inactive and damaged spores are removed, and the healthy spores are reserved for use; B: Put the substrate in the hole seedling tray, spray water to wet the surface of the substrate, inoculate the healthy spores in step A in the substrate, then sow sweet sorghum seeds on the substrate, and light culture for 40-50 days to obtain the culture; C: Take 1 / 4 of the whole culture vertically and wet sieve, check the amount of mycelium and spore propagation, discard the culture without infection and spores, and reserve the culture containing spores; D: Add substrate to the pot, move the culture containing spores into the pot, sow sweet sorghum seeds again, and continue to light culture for 3 months, then separate the spores to obtain spores after single spore culture.

3. The method for repairing cadmium contaminated soil by using arbuscular mycorrhizal fungi combined with sweet sorghum according to claim 2, characterized in that, The dark culture temperature in step (2) is 26°C, and the culture time is 20 days.

4. The method for repairing cadmium contaminated soil by using arbuscular mycorrhizal fungi combined with sweet sorghum according to claim 3, characterized in that, The preparation method of the accelerator is as follows: S1: Mix barium sulfate and dimethylformamide, stir at a speed of 500-800 r / min for 5-10 min to obtain a barium sulfate mixture, and reserve for use; S2: adding 60wt% ethanol solution into magnesium stearate and mixing uniformly, then placing in hot water to obtain magnesium stearate solution for use; S3: mixing maltose, L-methionine and wheat germ peptide, then adding water to stir and mix uniformly, then adding barium sulfate mixed solution and magnesium stearate solution, and placing in an ultrasonic homogenizer to obtain a promoter.

5. The method for repairing cadmium contaminated soil by using arbuscular mycorrhizal fungi combined with sweet sorghum according to claim 4, characterized in that, The temperature of the hot water for water bath heating in step S2 is 60-90℃, and the water bath heating time is 8-15min.

6. The method for repairing cadmium contaminated soil by using arbuscular mycorrhizal fungi combined with sweet sorghum according to claim 5, characterized in that, The homogenization conditions in step S3 are: frequency of 20-25kHz, temperature of 25-35℃, and homogenization time of 5-15min.

7. The method for repairing cadmium contaminated soil by using arbuscular mycorrhizal fungi combined with sweet sorghum according to claim 6, characterized in that, The application amount of the accelerator is 30-100 g / m 2 .

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