CN114854598B - Mortierella alpina and application thereof - Google Patents

Mortierella alpina and application thereof Download PDF

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CN114854598B
CN114854598B CN202210281574.5A CN202210281574A CN114854598B CN 114854598 B CN114854598 B CN 114854598B CN 202210281574 A CN202210281574 A CN 202210281574A CN 114854598 B CN114854598 B CN 114854598B
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陈林
张佳宝
李含放
赵炳梓
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    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
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Abstract

Mortierella alpina and application thereof, wherein the Mortierella alpina is preserved in a preservation unit designated by the national intellectual property agency, and the preservation date is 2021, 11 and 30, and the preservation unit name is as follows: china general microbiological culture Collection center, with the preservation number: CGMCC No.23826. The Mortierella alpina can be stably propagated in soil and can form a stable synergistic relationship with indigenous microorganisms. The mortierella alpina is derived from typical salty soil, and has the advantages of environmental protection, safety and high quality. The Mortierella alpina has the advantages of simple culture process, low preparation cost of the microbial inoculum and contribution to popularization.

Description

Mortierella alpina and application thereof
Technical Field
The invention belongs to the field of microorganism mortierella alpina, and particularly relates to mortierella alpina and application thereof.
Background
Fungi in the soil are mainly distributed on the surface layer of the soil, and among soil microorganism groups, the fungi are the main force for decomposing organic matters. Some beneficial fungi in the soil promote the mutual transformation of soil nutrients by stimulating the secretion of crop growth hormone, improve the stress resistance while improving the nutrient absorption of crops, and promote the growth of crops.
Mortierella alpinaMortierella alpina) Is a filamentous fungus of Mortierella genus of Mortierella order, and is widely distributed in soil, feces, and other spoilage organic materials. Due to the fact that the combination belongs to MortierellaMortierella) Mortierella pusilla can accumulate a considerable proportion of Arachidonic Acid (Arachidonic Acid, abbreviated)Called ARA or AA) has received a great deal of attention. Arachidonic acid, all-cis delta-5, 8,11, 14-eicosatetraenoic acid (ARA), is one of the three fatty acids necessary for human growth and development (Bajpai P K, bajpai P, ward O P Arachidonic acid production by fungi [ J ]]Appl Environ Microb,1991, 57:1255-1258.) belonging to the omega-6 series of polyunsaturated fatty acids (Cao just, convergence, liao Xiangru, cai Yujie. Fermentation and aging processes for producing arachidonic acid by Mortierella alpina [ J)]Food and biotechnology journal, 2016,35 (06): 617-622. Mortierella alpina is the only strain with formal safety evaluation in the current fungi for producing arachidonic acid, and the mycelium of the Mortierella alpina has high arachidonic acid content and is convenient to extract, so that the Mortierella alpina becomes a recognized excellent strain for producing arachidonic acid.
Mortierella alpina can accumulate polyunsaturated fatty acids such as Linoleic Acid (LA), linolenic acid (GlA), and eicosapentaenoic acid (EPA) in addition to arachidonic acid. In addition, in agriculture, mortierella alpina has been recently reported to play a role in degradation of organic materials, and studies have shown that mortierella elongata promotes carbon nitrogen phosphorus nutrient conversion of soil and secretion of plant growth hormone, thereby promoting plant growth and improving soil fertility (Li, f., chen, l., redstructure-Gordon, m., zhang, j.b., zhang, c.z., ning, q., li, w., 2018).Mortierella elongata's roles in organic agriculture and crop growth promotion in a mineral soil. Land Degradation &Development 29: 1642-1651). Meanwhile, research shows that the synergistic effect of the mortierella alpina and mortierella elongata mixed microbial inoculum obviously improves the quick-acting carbon and nitrogen nutrients and the related enzyme activities of carbon, nitrogen and phosphorus of soil, improves the soil fertility, and simultaneously promotes plants to secrete growth hormone so as to promote the growth of the plants (Ning Qi, chen Lin, li Fang, zhang Congzhi, ma Donghao, cai Zejiang, zhang Jiabao, 2020. The influence of mortierella on the soil nutrient availability and straw degradation. DOI: 10.11766/trxb 202006020213). However, the application of Mortierella alpina in saline soil is freshly reported at present, and the Mortierella alpina is isolated and cultured from the wheat rhizosphere saline soil of the yellow river delta research center of the national academy of sciences and resource researchMortierella alpina HSJ001）The Mortierella jenkinii preparation can effectively improve activity of enzymes related to carbon, nitrogen and phosphorus in saline soil, plays a certain role in activating carbon, nitrogen and phosphorus nutrients, can enable plant root systems to secrete a large amount of abscisic acid, promote nutrient absorption of plants, improve capability of the plants for coping with salt stress, and is beneficial to growth of the plants in saline soil environments. The Mortierella alpina can be prepared into a microbial inoculum and popularized and applied in the agricultural field.
Disclosure of Invention
The technical problems to be solved are as follows: the invention provides mortierella alpina and application thereof. The Mortierella alpina is obtained from wheat rhizosphere saline soil of yellow river delta research center of China academy of sciences and resource research, and is easy to culture and quick to reproduce. The strain has remarkable effect in promoting crop growth and activating soil nutrients. Meanwhile, the strain can promote the direct influence of the abscisic acid secreted by crops on the crop growth stress resistance.
The technical scheme is as follows: mortierella alpinaMortierella alpinaHSJ 001) which has been deposited in a deposit unit designated by the national intellectual property agency, the date of deposit being 2021, 11, 30, deposit unit name: china general microbiological culture Collection center, with the preservation number: CGMCC No.23826.
The method for culturing Mortierella alpina comprises the following steps: selecting Mortierella alpina filaments, inoculating the Mortierella alpina filaments into a PD liquid culture medium, and culturing the Mortierella alpina filaments at a constant temperature of 27 ℃ and 180 rpm for 7-10 days to obtain mycelium pellets; filtering under aseptic condition, adding 5 g mycelium pellets into 50 g wheat bran-bean cake powder-flour culture medium, and immediately shaking; standing at 27deg.C for culturing; after the mycelium solidifies the wheat bran-bean cake powder-flour culture medium into blocks, the two sides of the fungus cake can be contacted with air; then placing in a 27 ℃ incubator until the mycelium is completely covered with wheat bran-bean cake powder-flour culture medium, and mashing.
The PD liquid culture medium comprises the following components: peeled potato 200 g, glucose 20 g, water 1000 mL; the bran-bean cake powder-flour culture medium comprises the following components: wheat bran 15 g, bean cake powder 20 g, flour 15 g and water 50 mL, wherein the water is added with the bean cake powder to moisten for 30 min, then the wheat bran and the flour are added to be uniformly mixed, and the mixture is sterilized at 121 ℃ for 20 min and then uniformly shaken to obtain the bean cake.
The Mortierella alpina has the effect of promoting crop growth.
The application of the mortierella alpina in improving the activities of soil phosphatase, beta-glucosidase and N-acetyl-glucosaminidase.
The application of the mortierella alpina in synthesizing abscisic acid from plant root systems.
The mortierella alpina has the function of improving the dissolution of soil phosphorus.
The composition of the inorganic phosphorus culture medium for improving the dissolution of the soil phosphorus is as follows: glucose 10 g, (NH) 4 ) 2 SO 4 0.1 g、MgCl 2 ·6H 2 O 5.0 g、MgSO 4 ·7H 2 O 0.25 g、Ca 3 (PO 4 ) 2 5.0 g。
The beneficial effects are that: (1) The Mortierella alpina can improve the activity of alkaline phosphatase in soil (figure 1:a), remarkably improve the content of quick-acting phosphorus (figure 1:b), and has certain phosphorus dissolving capacity (figure 2), which shows that the Mortierella alpina can play a role in soil phosphorus conversion. (2) The mortierella alpina has the advantages that the activity of N-acetyl-glucosaminidase is obviously improved by 72.06 percent, the activity of beta-glucosaminidase is improved by 15.76 percent (shown in 1:a), which shows that the mortierella alpina plays an obvious role in activating soil nutrients, and the biomass of plants is obviously improved by 57.28 percent (shown in 1:f). (3) The content of abscisic acid in the root system of the crops is increased by 24.76 percent (figure 1:e), the content ratio of sodium to potassium in the leaves is reduced by 91.05 percent (figure 1:d), which shows that the strain plays a remarkable role in improving the salt tolerance and stress resistance of the crops. (4) The strain is used as rhizosphere microorganism, so that the biomass carbon content of rhizosphere soil microorganism is obviously improved by 89.80% (figure 1:c), and meanwhile, the microbial community structure is changed to regulate the crop growth (figure 3).
Drawings
FIG. 1 is a graph showing the effect of inoculation of Mortierella alpina on soil carbon, nitrogen, phosphorus-related enzyme activity, soil microbial biomass carbon content and quick-acting phosphorus content, and on wheat plant biomass, root system abscisic acid content, and leaf sodium content to leaf potassium content ratio. Wherein a is the influence of Mortierella alpina on enzyme activity, b is the influence of Mortierella alpina on quick-acting phosphorus content, c is the influence of Mortierella alpina on biochar content, d is the influence of Mortierella alpina on the ratio of sodium content to potassium content of leaves, e is the influence of Mortierella alpina on the content of abscisic acid of wheat root system, and f is the influence of Mortierella alpina on biomass of wheat plants.
FIG. 2 shows the growth of Mortierella alpina on inorganic phosphorus medium and its phosphorus dissolution amount.
FIG. 3 is the effect of inoculation of Mortierella alpina on soil rhizosphere fungal communities.
Detailed Description
The following examples are given solely for the purpose of illustrating the general principles and features of the present invention and are not meant to limit the scope of the invention in any way, as will be apparent to those skilled in the art. All equivalent changes or modifications made according to the spirit of the present invention should be included in the scope of the present invention.
The Mortierella alpina can be stably propagated in soil and can form a stable synergistic relationship with indigenous microorganisms. The composition of the inorganic phosphorus culture medium for improving the soil phosphorus dissolution of the mortierella alpina is as follows: glucose 10 g, (NH) 4 ) 2 SO 4 0.1 g、MgCl 2 ·6H 2 O 5.0 g、MgSO 4 ·7H 2 O 0.25 g、Ca 3 (PO 4 ) 2 5.0 g. The mortierella alpina is derived from typical salty soil of yellow river delta, and has the advantages of environmental protection, safety and high quality. The Mortierella alpina has the advantages of simple culture process, low preparation cost of the microbial inoculum and contribution to popularization.
Example 1
Separating and culturing Mortierella alpina from wheat rhizosphere saline soil of yellow river delta research center of national academy of sciences of China and research instituteMortierella alpinaHSJ 001) the strain was observed under a microscope to have tubular hyphae, developed hyphae, dense growth, lotus node-like hyphae and apparent transparent rings in inorganic phosphorus medium. The strain is preserved in a preservation unit specified by the national intellectual property agency, wherein the preservation date is 2021, 11, 30 and the name of the preservation unit: in (a)The national institutes of microbiological culture collection center, address: beijing, chaoyang area, north Chen Xi Lu 1, 3, china academy of sciences microbiological institute, deposit number: CGMCC No.23826. The nutrient indexes of the surface soil are as follows: the salt content of the soil is 2.14g/kg, the pH value is 8.16, the conductivity is 1.00ds/m, the organic carbon content of the soil is 7.13 g/kg, and the total phosphorus content of the soil is 0.73 g/kg.
The implementation steps are as follows:
step 1: and inoculating the Mortierella alpina strain with good growth into the PD liquid culture medium, and performing shaking culture at the temperature of 27 ℃ and at the speed of 180 rpm for 7-10 days.
Step 2: filtering the liquid-cultured Mortierella alpina under aseptic condition, and washing the filtered mycelium with sterile deionized water.
Step 3: 3 g clean mycelium is dissolved in 30 mL sterile deionized water, bacterial liquid is added into 1.5 kg soil, and the mixture is evenly mixed and then is filled into a basin.
Step 4: and (5) planting crops after fully watering.
Comparative example 1
The implementation steps are as follows:
step 1: accurately measuring 30 mL sterile deionized water without mycelium, adding 1.5 kg soil, and then filling into a basin.
Step 2: and (5) planting crops after fully watering.
Example 2
The implementation steps are as follows:
step 1: and inoculating the Mortierella alpina strain with good growth into the PD liquid culture medium, and performing shake culture at 27 ℃ and 180 rpm for 7-10 days until the liquid culture medium is turbid and a large number of mycelium pellets grow.
Step 2: filtering the liquid-cultured Mortierella alpina under aseptic condition, and washing the filtered mycelium with sterile deionized water.
Step 3: inoculating clean mycelium into sterilized wheat bran-bean cake powder-flour culture medium, standing at 27deg.C, and culturing in dark until mycelium completely covers the culture medium.
Step 4: and (3) mashing wheat bran, bean cake powder and flour culture medium of hyphae in different triangular flasks, taking out, and uniformly mixing to obtain the microbial inoculum.
Step 5: accurately weighing 3 g microbial inoculum, adding into 1.5 kg soil, fully mixing, and filling into a basin.
Step 6: and (5) planting crops after fully watering.
Comparative example 2
The implementation steps are as follows:
step 1: accurately weighing sterilized wheat bran-bean cake powder-flour culture medium 3 g without mycelium, adding into 1.5 kg soil, mixing thoroughly, and packaging in basin.
Step 2: and (5) planting crops after fully watering.
The soil to be tested is saline soil of a Shandong east camping farmland, and the nutrient indexes of the surface soil are as follows: the salt content of the soil is 3.50 g/kg, the organic carbon of the soil is 11.46 g/kg, the total nitrogen of the soil is 1.15 g/kg, the total phosphorus of the soil is 1.07 g/kg, and the quick-acting phosphorus of the soil is 32.07 mg/kg. The planted crop is wheat (Jinan 177), and three plants are reserved in each pot in the three-leaf period. Wheat is cultivated in a greenhouse environment, during which the moisture content is kept at 60%. And (5) watering and weeding are carried out according to the need in the period. Culturing to flowering period, sampling, and measuring soil enzyme activity, wheat biomass, and wheat root hormone content.
As shown in the measurement result in FIG. 1, the biomass of wheat treated by inoculation of Mortierella alpina is increased by 57.28%, the content of abscisic acid in the root system of wheat is increased by 24.76%, the ratio of sodium content in leaves to potassium content in leaves is reduced by 91.05%, the activity of alkaline phosphatase is increased by 58.04%, the activity of N-acetyl-glucosaminidase is obviously increased by 72.06%, and the activity of beta-glucosaminidase is increased by 15.76%. The result shows that the mortierella alpina can improve the secretion of abscisic acid of crops, and is favorable for the absorption of potassium ions by plant leaves, thereby improving the salt tolerance of crops. The increase of the beta-glucosidase activity, the phosphatase activity, the N-acetyl-glucosaminidase activity, the available phosphorus content and the wheat biomass of the soil treated by the inoculation of the mortierella alpina shows that the microbial inoculum has a promoting effect in the carbon, nitrogen and phosphorus activation of nutrients of saline soil, in particular to nitrogen and phosphorus nutrients, thereby improving the acquisition of the nutrients of crops and promoting the growth of the crops.

Claims (7)

1. A first partMortierella alpinaMortierella alpina) HSJ001, which has been deposited in a deposit unit designated by the national intellectual property agency, with a date of deposit of 2021, 11, 30, deposit unit name: china general microbiological culture Collection center, with the preservation number: CGMCC No.23826.
2. The method for culturing mortierella alpina according to claim 1, which is characterized by comprising the following steps: selecting Mortierella alpina filaments, inoculating the Mortierella alpina filaments into a PD liquid culture medium, and culturing the Mortierella alpina filaments at a constant temperature of 27 ℃ and 180 rpm for 7-10 days to obtain mycelium pellets; filtering under aseptic condition, adding 5 g mycelium pellets into 50 g wheat bran-bean cake powder-flour culture medium, and immediately shaking; standing at 27deg.C for culturing; after the mycelium solidifies the wheat bran-bean cake powder-flour culture medium into blocks, the two sides of the fungus cake can be contacted with air; then placing in a 27 ℃ incubator until the mycelium is completely covered with wheat bran-bean cake powder-flour culture medium, and mashing.
3. The method for culturing mortierella alpina according to claim 2, wherein the PD liquid medium comprises the following components: peeled potato 200 g, glucose 20 g, water 1000 mL; the bran-bean cake powder-flour culture medium comprises the following components: wheat bran 15 g, bean cake powder 20 g, flour 15 g and water 50 mL, wherein the water is added with the bean cake powder to moisten for 30 min, then the wheat bran and the flour are added to be uniformly mixed, and the mixture is sterilized at 121 ℃ for 20 min and then uniformly shaken to obtain the bean cake.
4. The use of mortierella alpina according to claim 1 for promoting crop growth.
5. Use of mortierella alpina according to claim 1 for increasing soil phosphatase, beta-glucosidase and N-acetyl-glucosaminidase activity.
6. The use of mortierella alpina according to claim 1 for synthesizing abscisic acid from plant roots.
7. The use of mortierella alpina according to claim 1 for increasing the solubilization of phosphorus in soil.
CN202210281574.5A 2022-03-21 2022-03-21 Mortierella alpina and application thereof Active CN114854598B (en)

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CN110747131B (en) * 2019-10-30 2020-12-29 中国科学院南京土壤研究所 Mortierella alpina and application thereof
CN110819539B (en) * 2019-11-27 2021-05-07 河南农业大学 Mortierella alpina 0901, application thereof and microbial inoculum
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