Sinorhizobium meliloti MCSWL1 and application thereof
By providing a MCSWL1 of Sinoorhizobium meliloti that can form a close symbiotic relationship with alfalfa, the problem of insufficient growth and yield of alfalfa in the prior art is solved, efficient nitrogen fixation and soil fertility improvement are achieved, and a wide range of agricultural application prospects are provided.
Patent Information
- Application Number
- CN202510002732.2
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-01-02
- Publication Date
- 2025-04-18
AI Technical Summary
The prior art is difficult to effectively improve the growth and yield of alfalfa, especially in the relationship between soil fertility and microbial symbiotic relationship.
A MCSWL1 of Sinoorhizobium meliloti is provided, obtained by screening from the rhizosphere of alfalfa and obtained by specific culture and preservation methods. This strain can form a close symbiotic relationship with alfalfa, perform efficient nitrogen fixation, and promote the growth of alfalfa.
The strain significantly increases alfalfa growth and yield, and increases soil fertility through a continuous supply of nitrogen, reduces fertilizer use, reduces agricultural production costs, and helps protect the ecological environment.
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Abstract
Description
Technical Field
[0001] The invention relates to the technical field of agriculture, and in particular to Sinorhizobium alfalfa MCSWL1 and applications thereof. Background Art
[0002] Alfalfa (Medicago sativa), as an important legume forage, is widely cultivated around the world, especially in animal husbandry. It is not only rich in high-quality protein and minerals, but also has good nitrogen fixation ability, which can significantly improve soil fertility. However, the growth and yield of alfalfa are affected by many factors, including soil fertility, climatic conditions, and microbial symbiosis.
[0003] Sinorhizobium meliloti, as an important type of rhizobium, has attracted widespread attention due to its wide host range and efficient nitrogen fixation ability. In recent years, with the increasing demand for sustainable agricultural development, there is an urgent need for efficient and stable Sinorhizobium meliloti resources to improve alfalfa production. Summary of the invention
[0004] In order to solve the above technical problems, the present invention provides a strain of Sinorhizobium meliloti, which exhibits a significant effect in promoting the growth of alfalfa, as follows: A Sinorhizobium meliloti MCSWL1, whose deposit number is CCTCC NO:M20231120, whose 16S rDNA sequence is shown in SEQ ID NO:1, and whose deposit date is June 28, 2023; The preservation name of the Sinorhizobium meliloti MCSWL1 is Sinorhizobium meliloti MCSWL1; the preservation unit is China Center for Type Culture Collection; and the preservation address is on the campus of Wuhan University, No. 299, Bayi Road, Wuchang District, Wuhan City, Hubei Province.
[0005] Moreover, the Sinorhizobium meliloti MCSWL1 was isolated and screened from the rhizosphere of alfalfa, and the specific method is as follows: 1. Preparation and dilution of alfalfa root soil stock solution The alfalfa root soil was dissolved in sterile water to prepare a soil stock solution. The prepared soil stock solution was diluted to 10 -1 , 10 -2 , 10 -3 Serial dilution; 2. Soil stock solution inoculation culture Inoculate the diluted soil stock solution on YMA medium; 3. Strain Purification Select a single colony from the medium of the initial inoculation culture and streak it onto a new medium for strain purification; 4. Secondary purification of strains Repeatedly streak the purified strains in new culture medium to improve the purity of the strains; 5. Bacterial culture and identification Prepare YMA liquid culture medium, inoculate the purified strain into the liquid culture medium and culture it in a shaker for 1-2 days. The bacterial liquid can be used for strain sequencing and strain preservation; 6. Preservation of bacterial strains The main methods of preserving laboratory strains include glycerol preservation, test tube slant preservation, and freeze-drying preservation.
[0006] On the other hand, the present invention provides a use of Sinorhizobium meliloti MCSWL1 for promoting the growth of alfalfa.
[0007] Moreover, the alfalfa is Grassland No. 3.
[0008] Moreover, a specific method for applying Sinorhizobium meliloti MCSWL1 is as follows: after activating Sinorhizobium meliloti MCSWL1, culturing in liquid for 2-3 days, diluting the MCSWL1 by 3-5 times, and watering the MCSWL1 on the roots of alfalfa.
[0009] Moreover, the amount of watering the alfalfa roots is 1 L per mu of land, which is sprayed into the soil around the seedlings.
[0010] Compared with the prior art, the present invention has the following beneficial effects: 1. The Sinorhizobium meliloti MCSWL1 provided by the present invention can absorb nitrogen in the air and convert it into ammonia that can be used by plants, form nodules, and provide nitrogen nutrition for plants.
[0011] 2. The nitrogen fixation effect of Sinorhizobium meliloti MCSWL1 provided by the present invention is not limited to current crops. In field experiments, we found that the strain can also provide nitrogen for subsequent crops, thereby playing a role in fertilizing the fields. This continuous nitrogen supply helps to improve soil fertility, reduce the use of chemical fertilizers, reduce agricultural production costs, and also help protect the ecological environment.
[0012] 3. The present invention provides an application of Sinorhizobium meliloti MCSWL1 for promoting the growth of alfalfa (Grassland No. 3), and the specific method is: after activating Sinorhizobium meliloti MCSWL1, it is diluted 3-5 times after liquid culture for 2-3 days, and watered on the roots of alfalfa. Through this method, the MCSWL1 strain can form a close symbiotic relationship with the roots of alfalfa and carry out efficient nitrogen fixation therein. DETAILED DESCRIPTION
[0013] Example 1 Acquisition of MCSWL1 strain A Sinorhizobium meliloti MCSWL1, whose accession number is CCTCC NO:M20231120, and whose 16S rDNA sequence is shown in SEQ ID NO:1; the Sinorhizobium meliloti MCSWL1 is isolated and screened from the rhizosphere of alfalfa, and the details are as follows: 1. Preparation and dilution of alfalfa root soil stock solution The alfalfa root soil was dissolved in sterile water to prepare a soil stock solution. The prepared soil stock solution was diluted to 10 -1 , 10 -2 , 10 -3 Serial dilution; 2. Soil stock solution inoculation culture Inoculate the diluted soil stock solution on YMA medium; 3. Strain Purification Select a single colony from the medium of the initial inoculation culture and streak it onto a new medium for strain purification; 4. Secondary purification of strains Repeatedly streak the purified strains in new culture medium to improve the purity of the strains; 5. Bacterial culture and identification Prepare YMA liquid culture medium, inoculate the purified strain into the liquid culture medium and culture it in a shaker for 1-2 days. The bacterial liquid can be used for strain sequencing and strain preservation; 6. Preservation of bacterial strains The main methods of preserving laboratory strains include glycerol preservation, test tube slant preservation, and freeze-drying preservation.
[0014] Example 2 Application of MCSWL1 strain The Sinorhizobium meliloti MCSWL1 prepared in Example 1 was used to promote the growth of alfalfa.
[0015] Moreover, the alfalfa is Grassland No. 3.
[0016] The specific method is: after activating Sinorhizobium meliloti MCSWL1, dilute it 4 times after 4 days of liquid culture and irrigate the roots of alfalfa.
[0017] Moreover, the amount of watering the alfalfa roots is 1 L per mu of land, which is sprayed into the soil around the seedlings.
[0018] Example 3: Verification of nitrogen fixation ability of MCSWL1 1. Strain culture: After the MCSWL1 strain was activated, the activated strain was inoculated into YMA liquid culture medium under sterile conditions and cultured for 2-3 days until the strain reached the logarithmic growth phase.
[0019] 2. Bacterial solution dilution and watering: The cultured bacterial solution was diluted 3-5 times; Grassland No. 3 alfalfa seedlings (experimental group) with uniform growth were selected, and the diluted bacterial solution was sprayed on the soil around the seedlings at a dosage of 1L per mu of land.
[0020] 3. Control group setting: A group of grassland 3 alfalfa not inoculated with MCSWL1 strain was set as the control group, and the other culture conditions were the same as those of the experimental group.
[0021] 4. Training and management: The alfalfa seedlings in the experimental group and the control group were cultured under the same environmental conditions, and the growth of alfalfa, such as plant height, leaf area, biomass, etc., was regularly observed and recorded.
[0022] 5. Determination of nitrogen fixation capacity: After 30 days of cultivation, the grassland No. 3 alfalfa of the experimental group and the control group were collected, and the nitrogenase activity in the roots of the experimental group and the control group, as well as the growth of alfalfa, were determined by acetylene reduction method to evaluate the nitrogen fixation ability of the MCSWL1 strain. See Table 1 for details.
[0023] Table 1 Verification results of nitrogen fixation ability of MCSWL1 strain
[0024] Note: The data in the table are mean ± standard deviation, and 10 alfalfa seedlings were collected from each group for measurement. As can be seen from Table 1, the nitrogenase activity of Grassland No. 3 alfalfa inoculated with MCSWL1 strain in the experimental group was significantly higher than that in the control group. At the same time, the plant height, leaf area and biomass were also significantly improved, which indicates that the MCSWL1 strain of Sinorhizobium alfalfa has significant nitrogen fixation ability and can promote the growth of alfalfa.
[0025] Example 4 Effect of MCSWL1 strain on soil fertility The same farmland was divided into two identical experimental fields, one of which was planted with alfalfa (Grassland No. 3) and irrigated once every 20 days using the technical solution of Example 2 (experimental group); the other experimental field was used as a control group, planted with the same alfalfa, and irrigated only with tap water at the same time.
[0026] 60 days later, after the alfalfa was harvested, the soil of the two experimental fields was sampled and analyzed to determine indicators such as the nitrogen content and organic matter content in the soil.
[0027] Table 2 Effects of MCSWL1 strain on soil fertility
[0028] Note: The data in the table are mean ± standard deviation, and 5 soil samples were collected for analysis in each group. As can be seen from Table 2, the soil nitrogen content and organic matter content of the experimental group were significantly higher than those of the control group, which indicates that the MCSWL1 strain can not only promote the growth of alfalfa, but also significantly improve the fertility of the soil, thereby playing the role of fertilizing the field. This continuous supply of nitrogen and organic matter helps to reduce the use of chemical fertilizers, reduce agricultural production costs, and also help protect the ecological environment.
[0029] Example 5 Effect of MCSWL1 strain on subsequent crops In the two experimental fields after the harvest in Example 4, wheat was planted after the land was cleaned, and the subsequent crops were routinely managed with the same irrigation and fertilization conditions (tap water was used once in the first half of the month, and fertilizer was applied uniformly once every 30 days). The crops were harvested after 60 days, and the plant height, leaf area, biomass and yield of the subsequent wheat were measured; soil samples were collected from the experimental group and the control group, and the soil nitrogen content was determined. The results are shown in Table 3.
[0030] Table 3 Effects of MCSWL1 strain on subsequent crops
[0031] Note: The data in the table are mean ± standard deviation. Ten wheat plants and five soil samples were collected for analysis in each group. As can be seen from Table 3, the plant height, leaf area, biomass and yield of wheat in the experimental group were significantly better than those in the control group, which indicates that the nitrogen fixation of the MCSWL1 strain on the previous alfalfa not only promoted the growth of alfalfa, but also significantly improved the soil fertility, and had a positive effect on the growth of the subsequent wheat. At the same time, the soil nitrogen content of the experimental group was also significantly higher than that of the control group, further confirming that the MCSWL1 strain can continuously provide nitrogen to the soil, which helps to improve soil fertility and crop yield. These results show that the MCSWL1 strain has broad application prospects in agricultural production, can reduce the use of chemical fertilizers, reduce agricultural production costs, and also help protect the ecological environment.
Claims
1. A Sinorhizobium meliloti MCSWL1, characterized in that: Its deposit number is CCTCC NO:M20231120, and its 16S rDNA sequence is shown in SEQ ID NO:
1.
2. The Sinorhizobium meliloti MCSWL1 according to claim 1, characterized in that: The Sinorhizobium meliloti MCSWL1 is isolated and screened from the rhizosphere of alfalfa.
3. Use of Sinorhizobium meliloti MCSWL1 as claimed in claim 1 or 2.
4. The use of Sinorhizobium meliloti MCSWL1 as claimed in claim 3, characterized in that: Used to promote alfalfa growth.
5. The use of Sinorhizobium meliloti MCSWL1 as claimed in claim 4, characterized in that: The alfalfa is Grassland No.
3.
6. The use of Sinorhizobium meliloti MCSWL1 as claimed in claim 5, characterized in that: The specific method is: after activating Sinorhizobium meliloti MCSWL1, dilute it 3-5 times after 3-5 days of liquid culture, and irrigate the roots of alfalfa.
7. The use of Sinorhizobium meliloti MCSWL1 as claimed in claim 6, characterized in that: The amount of irrigation for the alfalfa roots is 1 L per mu of land, which is sprayed into the soil around the seedlings.