Application of mycose composition in stress resistance regulation and control of saline-alkali soil

Abstract

The invention discloses an application of a mycose composition in saline-alkali soil stress resistance regulation, and belongs to the technical field of plant growth regulation. The microbial flora is bacillus; the plant source extract is salvia miltiorrhiza and safflower compound polysaccharide. The biological preservation number of bacillus in the composition is CGMCC (China General Microbiological Culture Collection Center) No.25566, and the bacterial concentration is 8.5 * 10 < 5 > CFU / mL; the salvia miltiorrhiza and safflower compound polysaccharide is prepared by mixing salvia miltiorrhiza and safflower as raw materials according to the mass ratio of (10-1): (1-10) through polysaccharide extraction, and the concentration of the salvia miltiorrhiza and safflower compound polysaccharide is 1 mu g / mL. The bacillus with the biological preservation number of CGMCC (China General Microbiological Culture Collection Center) No.25566, which is screened from medicinal plant salvia miltiorrhiza coepiphytic microorganisms, is combined with salvia miltiorrhiza and safflower compound polysaccharide for use, so that the bacterium-sugar composition for regulating and controlling the moderate and severe saline-alkali resistance of crops is obtained, and the application of salvia miltiorrhiza endophyte and plant-derived polysaccharide is further developed.

Description

Technical Field

[0001] The invention relates to the technical field of plant growth regulation, and in particular to application of a fungus sugar composition in stress resistance regulation of saline-alkali land. Background Art

[0002] my country's coastal saline-alkali land is widely distributed, accounting for nearly 40% of the total area of ​​saline soil in my country. Coastal saline-alkali soil is mainly distributed in the eastern coastal areas of the North China Plain, such as Liaodong Bay, Bohai Bay, Laizhou Bay and the coastal areas of Jiangsu and Zhejiang. The annual precipitation in the coastal area is generally above 500 mm, and the groundwater resources are abundant, with huge potential for development and utilization. Due to the permanent intrusion of seawater in coastal areas, too many soluble salts accumulate in the soil surface or soil body, and most coastal areas are mainly chlorides. At present, the treatment of coastal saline-alkali soil includes planting salt-tolerant plants, freshwater leaching, chemical improvement, and biological improvement. Among them, chemical improvement and biological improvement are more studied. Chemical improvement is to use some acidic salt substances to improve the properties of saline-alkali land, reduce the pH of the soil, increase the cation exchange capacity of the soil, and reduce the salt content of the soil. Biological improvement is to introduce organisms or microorganisms to improve soil structure, increase organic matter, and promote salt degradation. The above-mentioned improvers are all added to saline-alkali land, and there are few reports on directly regulating crop salt and alkali tolerance.

[0003] According to the third national soil survey, the salinity classification and grading standards for soil salinity are as follows: the salinity of saline-alkali farmland in coastal areas is classified into the salt content of the 0~20 cm topsoil layer (g / kg): mild is 1~2, moderate is 2~4, severe is 4~6, and saline soil / alkaline soil>6. Calculated in ‰, soils with a salt content of 3~6‰ are moderately and severely saline-alkali soils. At present, most of the saline-alkali land improvements are concentrated in mild and moderate saline-alkali lands. The patent with application number CN202310461661.3 discloses a multifunctional endophyte of Salvia miltiorrhiza and its application. An endophyte with good anti-plant pathogenic fungi activity was screened from the symbiotic microorganisms of the medicinal plant Salvia miltiorrhiza; the bacterium can also be lower than the salt stress tolerance and is used to increase the corn yield in mild and moderate saline-alkali land. The strain is used on 3‰ saline-alkali land to increase corn yield, while for moderately and severely saline-alkali land, the salt content is generally greater than 3‰, and it is difficult to achieve salt-alkali tolerance in moderately and severely saline-alkali land using only strains. Therefore, it is necessary to develop a bacterial agent composition containing endophytes of Salvia miltiorrhiza to regulate crop tolerance to moderately and severely saline-alkali, and further develop the application of endophytes of Salvia miltiorrhiza. Summary of the invention

[0004] In view of the above-mentioned prior art, the purpose of the present invention is to provide an application of a bacterial sugar composition in regulating the resistance of saline-alkali land. The present invention uses the Bacillus with a biological preservation number of CGMCC No. 25566 screened from the symbiotic microorganisms of the medicinal plant Salvia miltiorrhiza in combination with the Salvia miltiorrhiza safflower complex polysaccharide to obtain a bacterial sugar composition for regulating the tolerance of crops to moderate and severe salt and alkali, further expanding the application of endophytic bacteria of Salvia miltiorrhiza and plant-derived polysaccharides.

[0005] To achieve the above object, the present invention adopts the following technical solution: The present invention provides an application of a bacterial sugar composition in the stress resistance regulation of saline-alkali land, wherein the bacterial sugar composition comprises microorganisms and plant-derived extracts; The microorganism is Bacillus; the plant-derived extract is Salvia miltiorrhiza and safflower complex polysaccharide.

[0006] Preferably, the biological deposit number of the Bacillus is CGMCC No.25566.

[0007] Preferably, the salvia miltiorrhiza and safflower composite polysaccharide is prepared by mixing salvia miltiorrhiza and safflower as raw materials in a mass ratio of (10-1): (1-10) and extracting the polysaccharide.

[0008] Preferably, the preparation method of the Salvia miltiorrhiza and safflower composite polysaccharide is: The salvia miltiorrhiza and safflower were crushed and sieved, deionized water was added for ultrasonic-assisted extraction, centrifuged and filtered to obtain a water extract, sevage reagent was added to the water extract to remove protein, ethanol was added to the upper aqueous phase for alcohol precipitation, the mixture was refrigerated and allowed to stand, filtered and freeze-dried to obtain a salvia miltiorrhiza and safflower composite polysaccharide.

[0009] Preferably, the mass ratio of Salvia miltiorrhiza to Carthamus tinctorius is 3:1.

[0010] Preferably, the concentration of the Bacillus is 8.5×10 5 CFU / mL.

[0011] Preferably, the concentration of the Salvia miltiorrhiza and safflower complex polysaccharide is 1 μg / mL.

[0012] Preferably, the salt content of the saline-alkali land is 3~6‰.

[0013] Preferably, the application includes increasing the net photosynthetic rate, transpiration rate, stomatal conductance, dry weight per plant and yield of crops in saline-alkali land, and reducing the intercellular CO2 concentration of crops in saline-alkali land.

[0014] Preferably, the fungus-sugar composition is sprayed on the leaves of crops grown in saline-alkali land at a spraying rate of 225-375 L hm -2 .

[0015] Beneficial effects of the present invention: (1) The present invention uses Bacillus with biological preservation number CGMCC No. 25566 screened from the symbiotic microorganisms of the medicinal plant Salvia miltiorrhiza in combination with Salvia miltiorrhiza and safflower complex polysaccharides to obtain a bacterial sugar composition for regulating crop tolerance to moderate to severe salt and alkali, further expanding the application of endophytic bacteria of Salvia miltiorrhiza and plant-derived polysaccharides.

[0016] (2) Compared with chemical synthetic agents, the salvia miltiorrhiza and safflower complex polysaccharide component in the fungus sugar composition of the present invention has the advantages of being safer and having fewer side effects. The fungus sugar composition of the present invention acts on the leaves of crops, which can effectively activate the plant's defense mechanism and alleviate crop damage caused by moderate to severe salt stress. The corn salt tolerance test showed that foliar spraying once at the large bell mouth stage and the tasseling stage can significantly improve the crop's salt tolerance: increase the net photosynthetic rate, transpiration rate, stomatal conductance, dry weight per plant and yield, and reduce the intercellular CO2 concentration of crops in saline-alkali land. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 : Dry weight of maize plant in different treatment groups. DETAILED DESCRIPTION

[0018] It should be noted that the following detailed descriptions are illustrative and are intended to provide further explanation of the present application. Unless otherwise specified, all technical and scientific terms used herein have the same meanings as those commonly understood by those skilled in the art to which the present application belongs.

[0019] As introduced in the background technology section, most of the current crop salt-alkali tolerance is achieved by adding various types of improvers to saline-alkali land, while there are few reports on applying to crops to regulate crop salt-alkali tolerance. Moreover, most reports on salt-alkali tolerance are for mild to moderate salt-alkali tolerance, while it is more difficult to achieve moderate to severe salt-alkali tolerance.

[0020] Based on this, the purpose of the present invention is to provide an application of a bacterial sugar composition in the regulation of stress resistance in saline-alkali land. The present invention compounded Bacillus with biological preservation number CGMCC No. 25566 and Danshen Safflower composite polysaccharide, and the concentration of Bacillus was 8.5×10 5 CFU / mL, the concentration of Danshen Safflower Polysaccharide is 1μg / mL; spray on the leaves of crops. Taking corn as an example, spraying the leaves once at the big bell stage and the tassel stage can significantly improve the salt tolerance of crops, and its effect is significantly higher than using Bacillus or Danshen Safflower Polysaccharide alone.

[0021] When crops are subjected to abiotic stresses such as high temperature, salinity, and drought, it is very easy to cause imbalance of active oxygen metabolism and cell membrane damage in the plant body, resulting in damage to the photosynthetic system and causing plant wilting. The present invention combines microbial agents with functional plant polysaccharides. Microorganisms improve crop stress resistance by secreting growth hormones and promoting nutrient absorption; polysaccharides effectively remove the accumulation of active oxygen free radicals caused by salinity stress and improve the salinity stress resistance of crops. The combination of agents and polysaccharides can greatly and effectively improve the salt and alkali resistance of crops and promote yield increase.

[0022] In order to enable those skilled in the art to more clearly understand the technical solution of the present application, the technical solution of the present application will be described in detail below in conjunction with specific embodiments.

[0023] Description: The biological deposit number of the Bacillus used in the present invention is CGMCC No. 25566, and the depository institution is the General Microbiological Center of China Microbiological Culture Collection Administration. The strain is recorded in the patent with application number CN202310461661.3, entitled A multifunctional endophytic bacterium of Salvia miltiorrhiza and its application.

[0024] According to the "Third National Soil Survey Soil Salinity Classification Standard" in the "Implementation Plan for Special Survey of Saline-Alkali Soils", the salinity classification and classification of saline-alkali farmland in coastal areas is based on the salt content of the 0~20 cm topsoil layer (g / kg): mild is 1~2, moderate is 2~4, severe is 4~6, and saline soil / alkaline soil> 6. Therefore, calculated in ‰, soil with a salt content of 3~6‰ is moderately and severely saline-alkali soil.

[0025] The test materials used in the examples of the present invention are all conventional test materials in the art and can be purchased through commercial channels.

[0026] Example 1: Preparation of Salvia miltiorrhiza and Safflower Composite Polysaccharide Take 3 g of Danshen and 1 g of Carthamus tinctorius, crush and pass through a 40-mesh sieve; add 400 g of deionized water; perform ultrasonic-assisted extraction of polysaccharides, the ultrasonic frequency is 30KHz, the power is 150 W, and the ultrasonic time is 30 min. After extraction, centrifuge and filter to obtain a water extract, and add sevage reagent (n-butanol: chloroform in a volume ratio of 1:4) to the water extract to remove protein. The mass ratio of water extract to sevage reagent is 4:1. Take the upper aqueous phase and add ethanol to make the final ethanol concentration about 70wt%, refrigerate and stand at 4℃, filter after 24 hours, and freeze-dry at -30℃ for 48 hours to obtain Danshen and Carthamus tinctorius composite polysaccharide.

[0027] Example 2: Cultivation of Bacillus Bacillus (CGMCC No. 25566) was inoculated into liquid PDA medium and cultured at 200 rpm and 30 °C for 3 days to obtain a bacterial concentration of 8.5 × 10 5 CFU / mL of bacterial solution.

[0028] Example 3 Take the bacterial solution prepared in Example 2, add the Salvia miltiorrhiza and safflower complex polysaccharide prepared in Example 1 to a concentration of 1 μg / mL, mix well to obtain a saline-alkali soil stress resistance regulating bacterial sugar composition.

[0029] Comparative Example The salvia miltiorrhiza and safflower composite polysaccharide prepared in Example 1 was added into water to a concentration of 1 μg / mL to obtain a saline-alkali soil stress resistance regulating polysaccharide.

[0030] Test example 1. Experimental Setup The field experiment was conducted in Yibang Agricultural Science and Technology Experimental Base, Kenli District, Dongying City, Shandong Province (37°5'N, 118°36'E), which belongs to the warm temperate monsoon climate, with four distinct seasons and sufficient sunlight. Soil samples were collected at the test site on June 15, 2022, and the soil pH was 7.63, with an average salt content of 4‰, which belongs to moderate to severe coastal saline-alkali land.

[0031] The experimental material is the corn variety Denghai 605, which is sown on June 15th every year with a planting density of 67,500 plants hm2. -2 The row spacing was 60 cm and the plant spacing was 24.5 cm. The fertilizer used in the experiment was active humic acid compound fertilizer (N:P2O5:K2O=26:11:14), and the application rate was 750 kg hm -2 , applied once before sowing. Finely prepare the land and make soil moisture before sowing, set seedlings at the 3-leaf stage to reach the designed density, irrigate according to weather conditions and soil moisture, and prevent and control pests and diseases in time; other cultivation and management measures are the same as local farmers' habits. When corn grows to the set growth period, spray the leaves with exogenous substances.

[0032] The experiment was divided into four groups: a water control group (denoted as CK), Example 3 group (denoted as T), Example 2 group (denoted as SNMB1), and Comparative Example 1 group (denoted as 3S1C).

[0033] The experiment was carried out by spraying the leaves of corn once at the tasseling stage (V9) and the tasseling stage (VT). The spraying volume was 300 L hm-1 using an electric sprayer. -2 No other chemical control agents were used during the growth period. The planting scale of each group was: 5 m long, 6 m wide, and an area of ​​30 m 2 .

[0034] 2. Measurement items and methods 2.1 Leaf photosynthetic performance 2.1.1 Leaf area index and leaf photosynthetic potential Each group was treated at the flowering stage (R1), milky stage (R3) and mature stage (R6). Five plants with consistent growth were selected in the field, and the length and width of all their green leaves were measured to calculate the leaf area index (LAI) and leaf photosynthetic potential (LAD). Leaf area per fully expanded leaf (cm 2 ) = leaf length (cm) × leaf width (cm) × 0.75; Leaf area of ​​unexpanded leaves (cm 2 ) = leaf length (cm) × leaf width (cm) × 0.5; Leaf area index (LAI) = (leaf area per plant × number of plants per unit land area) / unit land area. LAD=(LA2+LA1) / 2×(t2-t1); Where LA1 and LA2 are the leaf area index per unit land area in the time periods t1 and t2, respectively.

[0035] 2.1.2 Gas exchange parameters In the R1 and R3 stages, the CIRAS-Ⅱ photosynthetic instrument (PP Systems, USA) was used to monitor the growth of the plants at 1600 µmol m -2 s -1 The photosynthetic performance of ear leaves was determined under different light intensity conditions. Ten representative plants were selected from each plot to determine the photosynthetic performance of ear leaves: net photosynthetic rate of ear leaves (Pn, μmol m -2 s -1 ), transpiration rate (Tr, mmol m -2 s -1 )、Stomatal conductance(Gs,mmol m -2 s -1 ) and intercellular CO2 concentration (Ci, μmol mol -1 ).

[0036] 2.2 Dry matter accumulation Five plants with the same growth were selected for each treatment at the flowering stage (R1), milky stage (R3) and mature stage (R6). The aboveground parts were separated into kraft paper bags according to the stems (including leaf sheaths), leaves, female ears (cobs + grains), male ears and bracts, and placed in an oven at 105℃ for 30 min, then dried at 80℃ to constant weight, and the dry weight was weighed after cooling.

[0037] 2.3 Output and output components During the corn harvest period, a 5 m long × 3 row wide plot with uniform growth was selected for harvesting and yield measurement. The number of plants, ears and double ear rate per unit area were counted. After natural air drying, the number of ear rows, ears per hectare and 1000-grain weight (water content 14%) were counted indoors and the yield per hectare was converted.

[0038] Production (kg / hm 2 ) = number of ears per hectare × grain weight per ear × thousand-grain weight.

[0039] 3. Test results 3.1 Leaf photosynthetic performance The photosynthetic performance of leaves is shown in Tables 1 and 2.

[0040] Table 1 Leaf area index and photosynthetic potential According to Table 1, after spraying T, SNMB1, and 3S1C, the leaf area index of the R1 stage increased by 22.8%, 9.3%, and 11.4% compared with CK; the leaf area index of the R3 stage increased by 22.8%, 10.4%, and 12.0% respectively compared with CK; the leaf area index of the R6 stage increased by 27.2%, 12.9%, and 14.2% respectively compared with CK. It is worth noting that the leaf area index of the 3S1C group is slightly higher than that of the SNMB1 group, indicating that the effect of using polysaccharides alone is slightly better than that of using bacterial agents alone. It can be seen that the saline-alkali land stress-resistant regulating bacterial sugar composition prepared by foliar spraying of Example 3 significantly improves the leaf area index of summer corn under salt stress in saline-alkali land compared with using bacterial agents or polysaccharides alone, so that it maintains a higher green leaf area during the growth period.

[0041] Compared with CK, after spraying T, SNMB1, and 3S1C, the total photosynthetic potential during the growth period of R1, R3, and R6 increased by 23.88%, 10.23%, and 13.03%, respectively. The increase in leaf photosynthetic potential improves the photosynthetic performance of the plant, which is beneficial to the accumulation of biomass in the plant population. It shows that the saline-alkali land stress-resistant regulating bacterial sugar composition prepared by foliar spraying Example 3 significantly improves the accumulation of biomass in the plant population compared with the use of bacterial agents or polysaccharides alone.

[0042] Table 2 Leaf gas exchange parameters According to Table 2, compared with CK, after spraying T, SNMB1, and 3S1C, the net photosynthetic rate Pn, transpiration rate Tr, and stomatal conductance Gs were all improved. It is worth noting that the transpiration rate of the 3S1C group was higher than that of the T group in the R1 period, but it dropped significantly in the R3 period. After spraying T, SNMB1, and 3S1C, the intercellular CO2 concentration Ci of the leaves was reduced, among which the T group decreased most significantly. The overall performance of gas exchange parameters was T>3S1C>SNMB1>CK, and the 3S1C group was slightly higher than the SNMB1 group. It shows that the saline-alkali land stress resistance regulating bacteria sugar composition prepared by foliar spraying of Example 3 can significantly improve the Pn, Tr, and Gs of summer corn leaves in saline-alkali land, reduce Ci, and provide guarantees for biomass accumulation and yield increase.

[0043] 3.2 Dry matter accumulation The dry matter accumulation is shown in Table 3 and Figure 1 .

[0044] according to Figure 1 It can be seen that compared with CK, the dry weight of individual plants at R6 stage increased by 31.18%, 14.24% and 15.16% after spraying T, SNMB1 and 3S1C treatments, respectively.

[0045] Table 3 Dry matter weight of maize per plant under different treatments According to Table 3, compared with the CK group, the dry matter of corn from R1 to R6 was increased after spraying T, SNMB1, and 3S1C. Among them, the increase in the T group was significantly higher than that in the SNMB1 group and the 3S1C group, indicating that the ability of adding the bacterial sugar composition to increase the dry matter of corn is significantly better than using SNMB1 bacteria alone or using Danshen Safflower complex polysaccharide alone. The bacterial sugar composition of the present invention is conducive to the accumulation of dry matter.

[0046] 3.3 Output and output components The corn yield and yield components of different treatment groups are shown in Table 4.

[0047] Table 4 Corn yield and its components in different treatment groups According to Table 4, the yield of T treatment is 11651kg / hm2 compared with CK. 2 , an increase of 18.34% compared with CK; the yield of SNMB1 treatment was 10603kg / hm 2 , an increase of 7.69% compared with CK; the yield of 3S1C treatment was 10710kg / hm 2, an increase of 8.78% over CK. Example 3 The yield of corn tolerant to moderate to severe saline-alkali can be significantly increased by using the bacterial agent + polysaccharide method, indicating that the bacterial sugar composition of the present invention can significantly improve the ability of corn to tolerate moderate to severe saline-alkali.

[0048] The above description is only the preferred embodiment of the present application and is not intended to limit the present application. For those skilled in the art, the present application may have various modifications and variations. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present application shall be included in the protection scope of the present application.

Claims

1. An application of a bacterial sugar composition in regulating stress resistance in saline-alkali land, characterized in that: The bacterial sugar composition comprises microorganisms and plant-derived extracts; The microorganism is Bacillus; the plant-derived extract is Salvia miltiorrhiza and safflower complex polysaccharide.

2. The use according to claim 1, characterized in that: The biological preservation number of the Bacillus is CGMCC No.25566.

3. The use according to claim 1, characterized in that: The salvia miltiorrhiza and safflower composite polysaccharide is prepared by mixing salvia miltiorrhiza and safflower as raw materials in a mass ratio of (10-1): (1-10) and extracting the polysaccharide.

4. The use according to claim 3, characterized in that: The preparation method of the salvia miltiorrhiza and safflower composite polysaccharide is as follows: The salvia miltiorrhiza and safflower were crushed and sieved, deionized water was added for ultrasonic-assisted extraction, centrifuged and filtered to obtain a water extract, sevage reagent was added to the water extract to remove protein, ethanol was added to the upper aqueous phase for alcohol precipitation, the mixture was refrigerated and allowed to stand, filtered and freeze-dried to obtain a salvia miltiorrhiza and safflower composite polysaccharide.

5. The use according to claim 4, characterized in that: The mass ratio of Salvia miltiorrhiza to Carthamus tinctorius is 3:

1.

6. The use according to claim 1, characterized in that: The concentration of the Bacillus was 8.5×10 5 CFU / mL.

7. The use according to claim 1, characterized in that: The concentration of the salvia miltiorrhiza and safflower complex polysaccharide is 1 μg / mL.

8. The use according to claim 1, characterized in that: The salt content of the saline-alkali land is 3~6‰.

9. The use according to claim 1, characterized in that: The application includes increasing the net photosynthetic rate, transpiration rate, stomatal conductance, dry weight per plant and yield of crops in saline-alkali land, and reducing the intercellular CO2 concentration of crops in saline-alkali land.

10. The use according to claim 1, characterized in that: Spray the fungus-sugar composition on the leaves of crops grown in saline-alkali land at a rate of 225-375 L hm -2 .

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