Method for measuring saline-alkaline tolerance pH threshold value of medicago sativa

Abstract

The invention relates to the technical field of agricultural ecology and crop cultivation, in particular to a method for measuring the saline-alkaline tolerance pH threshold value of medicago sativa L. The method comprises the following steps that black soil and soda saline-alkali soil are mixed in a gradient mode according to the mass ratio of (3-8): (2-7), and a soil environment with the pH being 8.64-9.91 is constructed; the alfalfa variety is sown in the soil environment, and the saline-alkaline tolerance pH threshold value of the alfalfa is determined according to the correlation between the plant height, the leaf number and the dry weight of the overground part of the alfalfa and the soil pH. According to the method disclosed by the invention, the saline-alkaline tolerance pH threshold value of alfalfa growth is determined to be 9.07 for the first time, and the suitable growth pH value is 8.60. In combination with field split region random experiment verification, the threshold value is stable and reliable in performance under field conditions, a scientific basis is provided for variety screening and soil improvement of saline-alkali soil alfalfa planting, and the method has the characteristics of simplicity and convenience in operation and reliable result.

Description

Technical Field

[0001] This invention relates to the fields of agricultural ecology and crop cultivation technology, specifically to a method for determining the pH threshold for salt and alkali tolerance in alfalfa. Background Technology

[0002] Soil salinization is a significant environmental problem restricting agricultural production. Soda-saline soils, in particular, with their high pH values ​​(typically >8.5), significantly inhibit plant growth, severely impacting crop yield and quality. Alfalfa (…) Medicago sativa As a high-quality legume forage, the quality of L.'s salt and alkali tolerance directly affects the sustainability and economic benefits of forage production in saline-alkali areas.

[0003] Traditional experimental methods often employ single saline-alkali soils or adjust pH using chemical reagents. These simplified treatments cannot realistically simulate the complex stress environment of soil salinization under natural conditions. Secondly, existing research lacks systematic quantitative analysis of key alfalfa growth indicators, such as plant height, leaf number, and biomass, in relation to soil pH gradient changes, making it difficult to accurately define their salt tolerance threshold. Furthermore, most variety screening trials remain at the laboratory stage, failing to incorporate confirmatory studies under field conditions, thus casting doubt on the reliability of the experimental results for practical application. These technical bottlenecks severely restrict the breeding of salt-tolerant alfalfa varieties and their widespread application in saline-alkali soils. Summary of the Invention

[0004] To address the above problems, this invention provides a method for determining the pH threshold of salt and alkali tolerance in alfalfa.

[0005] This invention is achieved through the following technical solution: A method for determining the pH threshold for salt and alkali tolerance in alfalfa, comprising the following steps: Black soil and soda saline-alkali soil were mixed in a gradient of 3-8:2-7 by mass to create a soil environment with a pH of 8.64-9.91. Several alfalfa varieties were sown in the soil environment described above. Plant height, number of leaves, and aboveground dry weight were measured during the 4th to 6th week of alfalfa growth. Plant height, number of leaves, and aboveground dry weight were measured again during the 7th to 9th week of alfalfa growth. The pH threshold for salt and alkali tolerance of alfalfa was determined by analyzing the correlation between the plant height, number of leaves, aboveground dry weight measured twice and soil pH.

[0006] Preferably, the black soil of the Nenjiang Riverbed and the soda saline-alkali soil are mixed in gradients at mass ratios of 8:2, 7:3, 5:5, 4:6 and 3:7, respectively.

[0007] Preferably, the black soil comes from the Nenjiang Riverbed.

[0008] Preferably, the alfalfa varieties are Bara218, Algunjin, Polar Bear, Bingchi, Gongnong NO.1, AlfaQueen, Aurora, GoldenEmpress, Optimus, Werner, Caribou, Chongjibo, Jingneng5010, Ladinuo, Longwei6010, Maitewei, Relang, Youxia, Gannong NO.1, Gannong NO.3, Gannong NO.6, Gannong NO.7, Gannong NO.8, and Magnum. NO.7, Longdong, MF4020, WL168HQ, WL298HQ, WL319HQ, WL343HQ, WL353HQ, WL354HQ, WL363HQ, WL366HQ and WL525HQ are any one or more of these.

[0009] Preferably, the alfalfa variety is sown in rows.

[0010] Preferably, the sowing amount for each alfalfa variety is 8g / row to 10g / row, and the row spacing is 20cm to 30cm.

[0011] Preferably, the method for determining plant height is to measure the straightened plant height of alfalfa seedlings.

[0012] Preferably, the method for determining the number of leaves is to count the total number of leaves per alfalfa seedling.

[0013] Preferably, the method for determining the aboveground dry weight is to blanch alfalfa seedlings at 107℃~109℃ for 29min~31min, dry them at 74℃~76℃ for 47h~49h, and then determine the aboveground dry weight.

[0014] Compared with the prior art, the present invention has the following beneficial effects: This invention provides a method for determining the pH threshold for salt and alkali tolerance of alfalfa, comprising the following steps: mixing black soil and soda saline-alkali soil in a gradient of 3-8:2-7 by mass to construct a soil environment with a pH of 8.64-9.91; sowing several alfalfa varieties in the soil environment; measuring plant height, number of leaves, and aboveground dry weight of alfalfa during the 4th to 6th week of growth; measuring plant height, number of leaves, and aboveground dry weight of alfalfa again during the 7th to 9th week of growth; and determining the pH threshold for salt and alkali tolerance of alfalfa by analyzing the correlation between the plant height, number of leaves, aboveground dry weight measured in the two measurements and the soil pH. This method constructs a soil environment with a pH range of 8.64–9.91 by mixing black soil from the Nenjiang Riverbed (pH 7.61) with soda saline-alkali soil at mass ratios of 8:2 to 3:7. Changes in plant height, leaf number, and individual plant biomass were measured for 35 alfalfa varieties. For the first time, the method clearly identifies alfalfa's pH threshold for salt and alkali tolerance as 9.07, with an optimal growth pH of 8.60. Combined with randomized controlled field experiments, this invention provides a scientific basis for variety selection and soil improvement for alfalfa cultivation in saline-alkali land, and is characterized by its simple operation and reliable results. Attached Figure Description

[0015] 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 some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0016] Figure 1 This image shows the planting process of alfalfa in greenhouse soil prepared according to the present invention.

[0017] Figure 2 The image shows the planting of alfalfa in the field according to the present invention.

[0018] Figure 3 This invention illustrates the dynamic changes in plant height of different alfalfa varieties under varying salt and alkali stress. Figure 3 In the diagram, A represents the dynamic changes in plant height of different alfalfa varieties under different salt and alkali stresses in week 5; B represents the dynamic changes in plant height of different alfalfa varieties under different salt and alkali stresses in week 8; C is the statistical chart corresponding to A; and D is the statistical chart corresponding to B.

[0019] Figure 4 This invention illustrates the dynamic changes in leaf number of different alfalfa varieties under varying salt and alkali stress. Figure 4 In the diagram, A represents the dynamic change in the number of leaves of different alfalfa varieties under different salt and alkali stresses in week 5; B represents the dynamic change in the number of leaves of different alfalfa varieties under different salt and alkali stresses in week 8; C is the statistical chart corresponding to A; and D is the statistical chart corresponding to B.

[0020] Figure 5 This invention illustrates the dynamic changes in the dry weight of a single alfalfa plant under different salt and alkali stresses of various alfalfa varieties. Figure 5 middle, Figure 4 In the diagram, A represents the dynamic change of single plant dry weight of different alfalfa varieties under different salt and alkali stresses in week 5; B represents the dynamic change of single plant dry weight of different alfalfa varieties under different salt and alkali stresses in week 8; C is the statistical chart corresponding to A; and D is the statistical chart corresponding to B.

[0021] Figure 6 This invention relates to the relationship between soil pH and alfalfa plant height and leaf number, as well as the relationship between plant height and leaf number. Figure 6 In the diagram, A represents the relationship between soil pH and alfalfa plant height; B represents the relationship between soil pH and leaf number; and C represents the relationship between plant height and leaf number. Detailed Implementation

[0022] To facilitate understanding of the present invention, a more comprehensive description is provided below, along with preferred embodiments. However, the present invention can be implemented in many different forms and is not limited to the embodiments described herein. Rather, these embodiments are provided to provide a thorough and complete understanding of the disclosure of the present invention.

[0023] Unless otherwise defined, all technical and scientific terms used in this invention have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. The terminology used in this invention and in its specification is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention.

[0024] Soil gradient configuration: Using alluvial black soil from the Nenjiang Riverbed with a pH of 7.61 as the control group, it was mixed with severely soda saline-alkali soil at different mass ratios to form gradient treatment groups with pH of 8.64 (mass ratio 8:2), pH of 9.07 (mass ratio 7:3), pH of 9.52 (mass ratio 5:5), pH of 9.7 (mass ratio 4:6), and pH of 9.91 (mass ratio 3:7).

[0025] The mixed soil settles naturally, simulating the physicochemical properties of real saline-alkali soil.

[0026] Determination of pH threshold for salt and alkali tolerance: Thirty-five alfalfa varieties were planted, and plant height, number of leaves, and biomass per plant were measured after 5 and 8 weeks.

[0027] Thresholds were determined by significant changes in growth indicators. Suitable pH range: pH≤8.60, with a slight increase in leaf number and biomass.

[0028] Tolerance limit threshold: at pH 9.07, plant height, number of leaves, and biomass decreased significantly.

[0029] Daejeon Verification: A split-plot completely randomized block design was used to verify the growth adaptability of alfalfa varieties at pH 8.60.

[0030] The beneficial effects of the present invention will be illustrated below through specific embodiments.

[0031] Example 1: A method for determining the pH threshold of salt and alkali tolerance in alfalfa Rectangular cement troughs are set up in parallel inside the greenhouse. The cement troughs are 7.0m long, 0.7m wide, and 0.6m high. The troughs are separated from each other by cement walls that are 7.0m long, 0.7m wide, and 0.1m high. The bottoms are sealed with cement to prevent water and salt migration between the soil in the troughs and to the outside environment.

[0032] The tested soils are shown in Table 2. Alluvial black soil from the Nenjiang Riverbed and severely soda-alkali soil from the experimental station area were used. The black soil had a pH of 7.61, while the soda-alkali soil had a pH > 10.0. Using black soil as a control group, black soil and soda-alkali soil were mixed at mass ratios of 8:2, 7:3, 5:5, 4:6, and 3:7, and then filled into different cement tanks. The total amount of black soil and soda-alkali soil in each tank was 3.5 tons. After natural settling for 29 days, test tanks with different pH gradients were formed, resulting in alkaline soil test treatment tanks with pH gradients of 8.64, 9.07, 9.52, 9.70, and 9.91, respectively.

[0033] Thirty-five alfalfa varieties were sown in each alkaline soil test treatment trench, with a sowing rate of 8g per row and a row spacing of 20cm. After emergence, weeding was carried out manually periodically, followed by normal management. An automatic temperature control system was used in the greenhouse: windows were opened when the temperature was ≥28℃ and closed when the temperature was ≤15℃. An automatic sprinkler system was in place to maintain light and humidity levels similar to outdoor conditions. Images of alfalfa planting in greenhouse soil preparation are shown below. Figure 1 As shown.

[0034] At weeks 4 and 7 of alfalfa growth, five representative plants of uniform growth from each variety were randomly selected. Their straightened plant height (cm) and total number of leaves per plant were measured. Twenty seedlings from each variety were randomly collected, blanched at 107℃ for 29 min, and then dried at 74℃ for 47 h. The dried biomass was measured and divided by 20 to obtain the dry weight per plant. The significant relationship between alfalfa plant height, leaf number, and aboveground dry weight and soil pH was statistically analyzed to determine the pH threshold for salt and alkali tolerance in alfalfa.

[0035] Example 2: A method for determining the pH threshold of salt and alkali tolerance in alfalfa Rectangular cement troughs are set up in parallel inside the greenhouse. The cement troughs are 8.0m long, 0.8m wide, and 0.7m high. The troughs are separated from each other by cement walls that are 8.0m long, 0.8m wide, and 0.7m high. The bottoms are sealed with cement to prevent water and salt migration between the soil in the troughs and to the outside environment.

[0036] The tested soils are shown in Table 2. Alluvial black soil from the Nenjiang Riverbed and severely soda-alkali soil from the experimental station area were used. The black soil had a pH of 7.61, while the soda-alkali soil had a pH > 10.0. Using black soil as a control group, black soil and soda-alkali soil were mixed at mass ratios of 8:2, 7:3, 5:5, 4:6, and 3:7, and then filled into different cement tanks. The total amount of black soil and soda-alkali soil in each tank was 3.75 tons. After natural settling for 30 days, test tanks with different pH gradients were formed, resulting in alkaline soil test treatment tanks with pH gradients of 8.64, 9.07, 9.52, 9.70, and 9.91, respectively.

[0037] Thirty-five alfalfa varieties were sown in each alkaline soil test treatment trough. The sowing rate for each alfalfa variety was 9g / row, with a row spacing of 25cm. After emergence, weeding was carried out manually periodically, followed by normal management. An automatic temperature control system was used in the greenhouse: windows were opened when the temperature was ≥28℃ and closed when the temperature was ≤15℃. An automatic sprinkler system was in place to maintain light and humidity levels similar to outdoor conditions. Images of alfalfa planting in greenhouse soil preparation are shown below. Figure 1 As shown.

[0038] At weeks 5 and 8 of alfalfa growth, five representative plants of uniform growth from each variety were randomly selected. Their straightened plant height (cm) and total number of leaves per plant were measured. Twenty seedlings from each variety were randomly collected, blanched at 108℃ for 30 min, and then dried at 75℃ for 48 h. The dried biomass was measured and divided by 20 to obtain the dry weight per plant. The significant relationship between alfalfa plant height, leaf number, and aboveground dry weight and soil pH was statistically analyzed to determine the pH threshold for salt and alkali tolerance in alfalfa.

[0039] Example 3: A method for determining the pH threshold of salt and alkali tolerance in alfalfa Rectangular cement troughs are set up in parallel inside the greenhouse. The cement troughs are 10.0m long, 1.0m wide, and 1.0m high. The troughs are separated from each other by cement walls that are 10m long, 1.0m wide, and 1.0m high. The bottoms are sealed with cement to prevent water and salt migration between the soil in the troughs and to the outside environment.

[0040] The tested soils are shown in Table 2. Alluvial black soil from the Nenjiang Riverbed and severely soda-alkali soil from the experimental station area were used. The black soil had a pH of 7.61, while the soda-alkali soil had a pH > 10.0. Using black soil as a control group, black soil and soda-alkali soil were mixed at mass ratios of 8:2, 7:3, 5:5, 4:6, and 3:7, and then filled into different cement tanks. The total amount of black soil and soda-alkali soil in each tank was 4.0 tons. After natural settling for 31 days, test tanks with different pH gradients were formed, resulting in alkaline soil test treatment tanks with pH gradients of 8.64, 9.07, 9.52, 9.70, and 9.91, respectively.

[0041] Thirty-five alfalfa varieties were sown in each alkaline soil test treatment trough, with a sowing rate of 10g per row and a row spacing of 30cm. After emergence, weeding was carried out manually periodically, followed by normal management. An automatic temperature control system was used in the greenhouse: windows were opened when the temperature was ≥28℃ and closed when the temperature was ≤15℃. An automatic sprinkler system was in place to maintain light and humidity levels similar to outdoor conditions. Images of alfalfa planting in greenhouse soil preparation are shown below. Figure 1 As shown.

[0042] At weeks 6 and 9 of alfalfa growth, five representative plants of uniform growth from each variety were randomly selected. Their straightened plant height (cm) and total number of leaves per plant were measured. Twenty seedlings from each variety were randomly collected, blanched at 109℃ for 31 min, and then dried at 76℃ for 49 h. The dried biomass was measured and divided by 20 to obtain the dry weight per plant. The significant relationship between alfalfa plant height, leaf number, and aboveground dry weight and soil pH was statistically analyzed to determine the pH threshold for salt and alkali tolerance in alfalfa.

[0043] Table 1 Test Materials

[0044] Note: " / " indicates that this item is not present.

[0045] Table 2 Basic physicochemical properties of the tested soils

[0046] Taking Example 1 as an example, the results of the growth dynamic changes of alfalfa under different salt and alkali stresses are as follows.

[0047] from Figure 3 As shown in A and B, the plant height of alfalfa showed a similar trend in weeks 5 and 8. With pH ranging from 7.61 to 8.64, plant height increased with increasing soil pH for different varieties. However, at pH 9.07, the plant height of most varieties showed a significant decreasing trend, subsequently decreasing with further increases in pH. From the mean... Figure 3Studies C and D showed that, compared to the control, at a pH of 9.07, plant height at weeks 5 and 8 decreased by 41.17% and 31.65%, respectively. When pH > 9.07, alkaline stress significantly inhibited alfalfa growth; the higher the soil pH, the greater the inhibitory effect on plant height. Slightly alkaline soil with a pH of around 8.64 was more suitable for alfalfa growth.

[0048] Depend on Figure 4 As shown in A and B, the number of leaves per alfalfa plant exhibits a similar trend to plant height, initially increasing and then decreasing. During each growth stage, pH 8.64 maintains the highest number of leaves, after which the number of aboveground leaves decreases with increasing pH. At pH 9.07, the number of aboveground leaves in most alfalfa plants shows a significant decreasing trend. Figure 4 As shown in C and D, the number of leaves at weeks 5 and 8 decreased by 31.40% and 18.37% respectively compared to the control. When the pH is greater than 9.52, the number of alfalfa leaves does not increase with the extension of growth time. The higher the soil pH, the stronger the inhibitory effect on alfalfa leaf formation; the slightly alkaline environment of pH 8.64 promotes the production of new leaves in alfalfa.

[0049] Depend on Figure 5 It was found that the aboveground dry weight of alfalfa at different growth stages showed similar trends as did plant height and leaf number. Aboveground dry matter accumulation reached its maximum at pH 8.64. At pH 9.07, compared to the control, dry weight decreased by 41.17% and 46.94% at weeks 5 and 8, respectively. This indicates that a high pH environment (pH > 9.07) inhibited aboveground biomass accumulation. Figure 5 As shown in C and D, the inhibitory effect is stronger as pH increases, and soil with pH 8.64 is more suitable for alfalfa growth.

[0050] It should be noted that the statistical analysis results in Examples 2 and 3 are the same as those in Example 1, indicating that a high pH environment (pH>9.07) inhibits the accumulation of aboveground biomass, and soil with pH 8.64 is more suitable for alfalfa growth.

[0051] Example 4, Field Verification Saline-alkali farmland with a pH gradient of 8.4–9.3 was selected, divided into separate plots, and alkali-tolerant varieties selected through screening were randomly planted. The reliability of the pH threshold determined in this invention was verified by comparing the plots with traditional planting areas.

[0052] The field trial was conducted on June 30, 2017, with 35 varieties participating in a split-plot experiment. The experiment was carried out in a completely randomized block design in saline-alkali soil, with each variety planted in an area of ​​3m². 2 For plots 1.5m wide and 2.0m long, sow in raised beds at a rate of 10g / 3m. 2The process is as follows: Alfalfa seeds are sown at a depth of 2 cm. After sowing, irrigate with well water with an EC of 1.05 mS / cm and a pH of 7.52. Weeding is carried out manually as needed later. Each variety is sown in four plots, thus including four biological replicates. See the attached image for an example of alfalfa planting in the field. Figure 2 As shown.

[0053] Five weeks after the seedlings began to grow, their height and number of leaves were measured in the field, and soil samples were collected from each plot using the "5-point method" to measure their pH and EC.

[0054] Field experiments verified that alfalfa thrives best at pH 8.6, while plant height and leaf number significantly decreased at pH 9. Soil pH showed a significant negative correlation with both plant height and leaf number. P <0.05. Plant height and leaf number showed a highly significant positive correlation. P <0.01. For example... Figure 6 As shown.

[0055] In summary, this invention found that alfalfa plant height, leaf number, and aboveground dry weight exhibited similar trends at different growth stages. Specifically, at pH 9.07, alfalfa growth indicators were significantly lower than the control group, and the degree of growth inhibition increased with increasing pH. Alfalfa varieties were more suitable for growth in both greenhouse and field soils at a pH of around 8.6.

[0056] This invention provides a scientific basis for variety selection and soil improvement for alfalfa cultivation in saline-alkali land, and is characterized by its simple operation and reliable results.

[0057] The technical features of the above embodiments can be combined in any way. For the sake of brevity, not all possible combinations of the technical features in the above embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0058] The embodiments described above are merely illustrative of several implementations of the present invention, and while the descriptions are specific and detailed, they should not be construed as limiting the scope of the invention. Those skilled in the art can make various modifications and improvements without departing from the concept of the present invention, and these all fall within the protection scope of the present invention. Therefore, the protection scope of this invention should be determined by the appended claims.

Claims

1. A method for determining the pH threshold for salt and alkali tolerance in alfalfa, characterized in that, The method comprises the following steps: mixing black soil and soda saline soil in a mass ratio of 3-8:2-7 to build a soil environment with a pH of 8.64-9.91; seeding several alfalfa varieties in the soil environment; measuring the plant height, leaf number and aboveground dry weight of the alfalfa in the 4th-6th week of growth; measuring the plant height, leaf number and aboveground dry weight of the alfalfa in the 7th-9th week of growth; and determining the salt-tolerant pH threshold of the alfalfa through correlation analysis of the plant height, leaf number, aboveground dry weight and soil pH.

2. The method of claim 1, wherein, The black soil and the soda saline soil are mixed in a mass ratio of 8:2, 7:3, 5:5, 4:6 and 3:7, respectively.

3. The method of claim 1, wherein, The black soil is from the Nenjiang River bed.

4. The method of claim 1, wherein, The alfalfa varieties are any one or more of Bara218, Algunjin, Polar bear, Bingchi, Gongnong NO.1, AlfaQueen, Aurora, Golden Empress, Optimus, Werner, Caribou, Chongjibo, Jingneng5010, Ladinuo, Longwei6010, Maitewei, Relang, Youxia, Gannong NO.1, Gannong NO.3, Gannong NO.6, Gannong NO.7, Gannong NO.8, Magnum NO.7, Longdong, MF4020, WL168HQ, WL298HQ, WL319HQ, WL343HQ, WL353HQ, WL354HQ, WL363HQ, WL366HQ and WL525HQ.

5. The method of claim 1, wherein, The seeding mode of the alfalfa varieties is drill seeding.

6. The method of claim 1, wherein, The seeding amount of each alfalfa variety is 8-10 g / row, and the row spacing is 20-30 cm.

7. The method of claim 1, wherein, The plant height is measured by measuring the straightened plant height of the alfalfa seedlings.

8. The method of claim 1, wherein, The aboveground dry weight is measured by killing green the alfalfa seedlings at 107-109 ℃ for 29-31 min, drying at 74-76 ℃ for 47-49 h, and then measuring the aboveground dry weight.

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