Application of palmitic acid in promoting symbiosis between arbuscular mycorrhizal fungi and maize

Abstract

The application discloses application of palmitic acid in promoting symbiosis of arbuscular mycorrhizal fungi and corn, and relates to the technical field of soil microbial fertilizer. The arbuscular mycorrhizal fungi can reach 38% of the infection frequency of corn, 866.92% higher than that of the CK control group, i.e. 3.93%, and the symbiosis of the arbuscular mycorrhizal fungi and the corn is significantly promoted; and the 2mmol / L palmitic acid treatment has a significant growth promoting effect on the corn.

Description

Technical Field

[0001] This invention relates to the field of soil microbial fertilizer technology, and more specifically to the application of palmitic acid in promoting the symbiosis of arbuscular mycorrhizal fungi and maize. Background Technology

[0002] Arbuscular mycorrhizal fungi (AMF) are widely distributed in nature and can form symbiotic relationships (mycorrhizae) with more than 80% of terrestrial plants. Through this symbiotic relationship, AMF promotes the absorption of mineral nutrients from the soil. AMF facilitates the absorption and utilization of minerals such as nitrogen (N), phosphorus (P), and potassium (K), as well as micronutrients like phosphates and nitrogen. This symbiotic relationship helps plants absorb water and nutrients, improves their stress resistance, and also enhances soil quality.

[0003] Therefore, providing a promoter that can enhance arbuscular mycorrhizal fungi is of great significance for promoting crop growth by utilizing symbiotic effects. Summary of the Invention

[0004] To achieve the above objectives, the present invention adopts the following technical solution:

[0005] Application of palmitic acid in promoting symbiosis between arbuscular mycorrhizal fungi and maize.

[0006] Preferably, the concentration of palmitic acid is 2 mmol / L to 4 mmol / L.

[0007] More preferably, the concentration of palmitic acid is 2 mmol / L.

[0008] Preferably, the solvent for diluting palmitic acid is 1% ethanol.

[0009] Another objective of this invention is to provide a method for promoting symbiosis between arbuscular mycorrhizal fungi and maize, wherein palmitic acid is added exogenously after maize seedling emergence.

[0010] Preferably, the concentration of palmitic acid is 2 mmol / L to 4 mmol / L.

[0011] More preferably, the concentration of palmitic acid is 2 mmol / L.

[0012] Preferably, the palmitic acid is applied three times after the corn emerges, with each application 10 days apart.

[0013] Another object of the present invention is to provide a promoter for the symbiotic relationship between arbuscular mycorrhizal fungi and maize, said promoter comprising 2 mmol / L to 4 mmol / L palmitic acid.

[0014] Preferably, the promoter comprises 2 mmol / L palmitic acid.

[0015] More preferably, the accelerator consists of palmitic acid and 1% ethanol solvent.

[0016] Beneficial effects: The exogenous addition of palmitic acid in this invention can increase the infection frequency of arbuscular mycorrhizal fungi and maize to 38%, which is 866.92% higher than the infection frequency of 3.93% in the CK control group, significantly promoting the symbiosis between arbuscular mycorrhizal fungi and maize; and the 2 mmol / L palmitic acid treatment also has a significant growth-promoting effect on maize. Attached Figure Description

[0017] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on the provided drawings without creative effort.

[0018] Figure 1 Microscopic images of AMF in soil;

[0019] Figure 2 Phenotypic images of maize plants at different concentrations of exogenous fatty acids after 25 days;

[0020] Figure 3 Phenotypic images of maize plants at different concentrations of exogenous fatty acids after 45 days;

[0021] Figure 4 : Bar graph of aboveground dry weight of maize at different concentrations of exogenous fatty acids after 25 days;

[0022] Figure 5 : Bar graph of aboveground dry weight of maize at different concentrations of exogenous fatty acids after 45 days;

[0023] Figure 6 : Bar graph of dry weight of maize underground parts under different exogenous fatty acid concentrations after 25 days;

[0024] Figure 7 : Bar graph of dry weight of maize underground parts at different concentrations of exogenous fatty acids after 45 days

[0025] Figure 8 :Infection of maize roots under different concentrations of exogenous fatty acids after 25 days;

[0026] Figure 9 :Infection of maize roots under different concentrations of exogenous fatty acids after 45 days;

[0027] Figure 10 :Column graph of maize root infection under different concentrations of exogenous fatty acids after 25 days;

[0028] Figure 11 : Column graph of maize root infection at different concentrations of exogenous fatty acids after 45 days;

[0029] In the above figures, Z represents palmitic acid, YZ represents stearic acid, YM represents linolenic acid, YS represents oleic acid, and YY represents linoleic acid. Detailed Implementation

[0030] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0031] Example 1

[0032] 1. Solubility of exogenous fatty acid additives

[0033] The exogenous fatty acid additives involved in this invention—linoleic acid, linolenic acid, oleic acid, palmitic acid, and stearic acid—were all purchased from Shanghai Aladdin Biotechnology Co., Ltd.

[0034] Dissolution of exogenous fatty acid additives: Dissolve the exogenous fatty acids first with a small amount of anhydrous ethanol, and then dilute the ethanol with pure water to 1%.

[0035] 2. Soil background value measurement and microscopic examination of AMF in soil

[0036] Yunnan red soil was selected.

[0037] (1) Background value determination: After the soil is sieved through a 5mm sieve and mixed, relevant background physicochemical properties are determined. The main parameters to be determined are soil moisture content, total nitrogen, total phosphorus, total potassium, nitrate nitrogen, ammonia nitrogen and pH.

[0038] Soil pH was determined by pure water extraction and a pH meter. Soil moisture content was measured using the oven-drying method, a very simple method for determining soil moisture. The procedure is as follows: approximately 5g of soil is weighed, placed in an aluminum box, and then placed in an oven at 105℃ until dried. The standard for drying is that the weight remains constant between two consecutive weighings. The amount of water lost after drying is the soil moisture content. Ammonia nitrogen was extracted with potassium chloride and determined using the indophenol blue colorimetric method; nitrate nitrogen was extracted with potassium chloride and determined using ultraviolet spectrophotometry; available phosphorus was extracted with sodium bicarbonate and determined using the molybdenum antimony colorimetric method; available potassium was extracted with ammonium acetate and determined using the flame photometry method; total nitrogen was digested with sulfuric acid and determined using the sodium salicylate method; total phosphorus was digested with sulfuric acid and determined using the molybdenum antimony colorimetric method; and total potassium was digested with sulfuric acid and determined using the flame photometry method.

[0039] The soil's baseline physicochemical values, as determined by the above methods, are as follows: soil pH 5.4, moisture content 13.5%, total phosphorus 0.87 g / kg, total nitrogen 2.15 g / kg, available phosphorus 3.36 mg / kg, total potassium 1.35 g / kg, ammonium nitrogen 3.29 mg / kg, and nitrate nitrogen 4.24 mg / kg.

[0040] (2) Microscopic examination of AMF in soil: After obtaining the soil, weigh 10-20g of air-dried soil sample and place it in a beaker. Soak it in 100mL of pure water for 20-30min. After larger stones or debris have settled at the bottom of the beaker, quickly pour the suspension through a three-layer sieve with mesh sizes of 25, 65, and 250. Use a wash bottle to rinse the filter material from the bottom sieve into a 100mL centrifuge tube. Centrifuge at 3000rpm for 3min to remove surface debris and supernatant. Then, gently rinse the residue remaining on the sieve surface with a wash bottle and gently rinse it into a clean petri dish. After transferring to the petri dish, observe and count the number of spores in the petri dish under an electron microscope. The number of AMF spores in the soil sample was 85 / kg. See Appendix for their morphology. Figure 1 .

[0041] Example 2

[0042] 1. Corn potted plant cultivation

[0043] A pot experiment was conducted using the B73 maize variety as the planting material. The pot experiment included two concentration treatments (2 mmol / L and 4 mmol / L), plus five different exogenous fatty acid additives (oleic acid, linoleic acid, linolenic acid, palmitic acid, and stearic acid), and a negative control group was set up. Each treatment was repeated 12 times, for a total of 132 pots.

[0044] The experiment was conducted in the greenhouse of the Institute of Resource Plants, Yunnan University (24.51°N, 102.51°E). Potted experiments were conducted using 10cm × 9cm pots. Soil collected in Example 1 was sieved through a 5mm sieve and mixed thoroughly; 1.5kg of sieved soil was initially added to each pot. B73 maize seeds were sterilized and germinated indoors; plump seeds were selected for sowing, with one seed sown per pot. The exogenous additives used in the experiment were oleic acid, linoleic acid, linolenic acid, palmitic acid, and stearic acid. Exogenous fatty acids, weighed according to different concentrations, were dissolved in 1% ethanol and applied in batches to the treatment groups after maize transplanting; the negative control group received the 1% ethanol solution. The exogenous fatty acid solution was applied three times, with 100ml of the prepared fatty acid solution added to each pot each time. The application times were 1 day, 10 days, and 20 days after maize transplanting. Afterwards, cultivate under natural light conditions, watering every 3-5 days to maintain 70% humidity for plant growth. The same cultivation and management measures were applied to the corn plants during the planting process (see appendix for plant phenotype 25 days after transplanting). Figure 2See Appendix for plant phenotype 45 days after transplanting. Figure 3 ).

[0045] 2. Determination of dry weight of corn aboveground and underground parts

[0046] Sampling was conducted at 25 and 45 days after planting. For each sampling, three plants were randomly selected from each treatment for destructive sampling. The above-ground and underground parts of the corn plants were separated using scissors. The roots were then soaked in clean water to wash away the soil on the root surface. After washing, the roots were dried with absorbent paper, and secondary roots were removed. The above-ground and underground dry-fresh weights of the corn plants were measured. The above-ground and underground parts were placed in separate envelopes. The fresh plant samples were placed in an oven at 105℃ for 15 minutes to blanch, then dried at 70℃ until constant weight was achieved, and finally weighed. The above-ground and underground dry weights of the plants were recorded (see Appendix). Figure 4-7 ).

[0047] From the appendix Figure 4-7 It can be seen that, at 25 days after planting, after treatment with different concentrations of fatty acids, the dry weight of both the aboveground and belowground parts of maize showed an increasing trend under the 2 mmol / L palmitic acid treatment. The results at 45 days after planting showed the same trend as at 25 days. Therefore, it can be seen that, compared with other treatments, the 2 mmol / L palmitic acid treatment can significantly increase the dry weight of maize.

[0048] 3. Determination of symbiotic relationship between arbuscular mycorrhizal fungi and maize

[0049] Sampling was conducted on days 25 and 45 after planting. During sampling, the corn plants, along with their roots, were carefully removed from each pot. The roots were then washed, dried with filter paper, and secondary roots were removed. The roots were cut into approximately 1 cm long segments and soaked in FAA fixative for at least 4 hours. The root segments were then removed, rinsed with tap water, and a suitable amount of 10% KOH solution was added. The roots were heated in a 90°C water bath for 30 minutes, then the KOH solution was discarded. The roots were rinsed several times in tap water, drained, and acidified with 2% hydrochloric acid for 5 minutes. The acidification solution was discarded, and a suitable amount of 0.05% trypan blue staining solution was added. Staining was performed overnight at room temperature. The staining solution was discarded, and the roots were decolorized repeatedly with lactic acid glycerol decolorizing solution until most of the excess dye was removed. The resulting slides were then examined under a microscope.

[0050] The infection frequency of root segments was graded using the cross-multiplication method. After destaining, 30-50 root segments were randomly selected and prepared into slides, with 10 segments pressed onto each slide. The infection frequency was then determined under a 200× microscope using the cross-multiplication method (see appendix). Figure 8-11 ).

[0051] From the appendix Figure 8 -Appendix Figure 11It can be seen that the addition of different fatty acids all showed an increasing trend in the indicators of maize root infection, with palmitic acid showing the most significant effect. At a concentration of 2 mmol / L, the addition of palmitic acid resulted in an AMF infection frequency of 38%, while the infection frequency of the control group was only 3.93%, representing an increase of 866.92%. This demonstrates that the addition of 2 mmol / L palmitic acid can significantly promote the symbiosis between arbuscular mycorrhizal fungi and plants.

[0052] The above experiments show that palmitic acid can promote the symbiosis between arbuscular mycorrhizal fungi and maize. Furthermore, the infection rate gradually increases with the number of days of planting, and the symbiotic phenomenon becomes increasingly pronounced.

[0053] The various embodiments in this specification are described in a progressive manner, with each embodiment focusing on the differences from other embodiments. The same or similar parts between the various embodiments can be referred to each other.

[0054] The above description of the disclosed embodiments enables those skilled in the art to make or use the invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the invention. Therefore, the invention is not to be limited to the embodiments shown herein, but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. The application of palmitic acid in promoting symbiosis between arbuscular mycorrhizal fungi and maize, characterized in that, The palmitic acid was applied three times after the corn seedlings emerged, with each application 10 days apart; the concentration of palmitic acid was 2 mmol / L; as the number of days after planting increased, the infection rate gradually increased, and the symbiotic phenomenon became more and more obvious; moreover, the 2 mmol / L palmitic acid treatment had a significant growth-promoting effect on corn.

2. A method for promoting symbiosis between arbuscular mycorrhizal fungi and maize, characterized in that, Palmitic acid was added exogenously after corn emergence. Specifically, it was applied three times after corn emergence, with an interval of 10 days between each application. The concentration of palmitic acid was 2 mmol / L. As the number of days after planting increased, the infection rate gradually increased, and the symbiotic phenomenon became more obvious. Moreover, the 2 mmol / L palmitic acid treatment had a significant growth-promoting effect on corn.

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