Soybean salt tolerance identification method

Abstract

The invention discloses a soybean salt tolerance identification method, which comprises the steps of soybean sprout period salt tolerance identification, seedling period salt tolerance identification and calculation by using a comprehensive evaluation calculation model on the basis of sprout period and seedling period identification results, the salt tolerance level of the soybean in the seedling stage is determined according to the relative salt damage index of the germination rate, and the salt tolerance level of the soybean in the seedling stage is determined by cultivating soybean seedlings in a cultivation pot and performing salt treatment, and determining the salt tolerance level of the soybean in the seedling stage through visual inspection of the plant growth condition of the soybean and the number of damaged leaves. The method solves the problem that a traditional soybean salt tolerance identification method adopts a single period or a specific growth stage to measure the salt tolerance, which is one-sided.

Description

Technical Field

[0001] The invention relates to the technical field of soybean breeding, and in particular to a method for identifying soybean salt tolerance. Background Art

[0002] Soybeans are an important cash crop in my country, playing a crucial role in both consumer consumption and livestock and poultry production. However, my country's soybean self-sufficiency rate is low, less than 20% in 2021, with over 80% dependent on imports. my country has become the world's largest soybean importer, and heavy reliance on imports poses risks to food security. my country is rich in saline-alkali land resources, and vigorously developing soybean cultivation in saline-alkali land has become a key avenue for the development of the soybean industry. Soybeans are moderately salt-tolerant crops, and screening for salt-tolerant soybean germplasm and cultivating salt-tolerant varieties are crucial to ensuring the development of the soybean industry.

[0003] Research on soybean salt tolerance primarily relies on accurate identification and evaluation. Different identification methods yield different final assessments of salt tolerance across different materials. Current salt tolerance screening and identification methods still have some limitations. For example, measuring salt tolerance based solely on a single period or specific growth stage can be one-sided. Soybeans can vary in their response mechanisms and tolerance to salt at each stage, and a single metric cannot capture these complex variations and sometimes fails to fully reflect actual conditions.

[0004] On the other hand, traditional identification methods are inefficient and have limited accuracy. For example, the Petri dish method, commonly used during the bud stage, limits root growth space, mismatches seedling growth with natural conditions, and lacks simulation. Furthermore, local salt distribution is uneven, and screening large-scale resources such as mutant libraries and natural populations requires high consumable material costs. Salt tolerance assessment at the seedling stage often relies on morphological or physiological indicators, which are complex, technically demanding, costly, and time-consuming. Summary of the Invention

[0005] In order to overcome the defects of the existing technology, a method for identifying soybean salt tolerance is provided to solve the problem that the traditional soybean salt tolerance identification method uses a single period or a specific growth stage to measure salt tolerance, which is relatively one-sided.

[0006] To achieve the above object, a method for identifying salt tolerance of soybean is provided, including identification of salt tolerance at the germination stage and identification of salt tolerance at the seedling stage;

[0007] The identification of soybean salt tolerance during the germination stage comprises the following steps: fully soaking multiple sterilized germination papers in clean water and a 1.2% NaCl solution, respectively; evenly placing soybeans on two layers of moistened germination paper, and then covering the soybeans with another germination paper of the same size; rolling the three layers of paper with the soybeans into a roll and placing it vertically, maintaining humidity, so that the soybeans germinate; using the soybeans germinated in the germination paper soaked in clean water as a control group, and calculating the relative salt damage index of the soybeans germinated in the germination paper soaked in the NaCl solution based on the measured germination rate and variety germination rate of the soybeans; and determining the germination stage salt tolerance level of the soybeans based on the relative salt damage index of the soybeans.

[0008] The seedling stage salt tolerance assessment includes placing soybeans in a substrate in a cultivation pot; placing the cultivation pot in an artificial climate chamber; watering the soybeans in the cultivation pot with a 0.6% NaCl solution for 14 days after the first pair of true leaves of the soybeans unfold; and determining the seedling stage salt tolerance level of the soybeans based on visual inspection of plant growth and the number of damaged leaves.

[0009] Based on the weights of the soybean salt tolerance assessment at the germination stage and the soybean salt tolerance assessment at the seedling stage, the soybean salt tolerance grade is calculated using a comprehensive evaluation calculation model, and the comprehensive evaluation calculation model is:

[0010] N = X × a + Y × b;

[0011] Among them, N is the salt tolerance grade of the soybean, X is the salt tolerance grade of the soybean at the germination stage, a is the weight of the salt tolerance identification of the soybean at the germination stage, Y is the salt tolerance grade of the soybean at the seedling stage, and b is the weight of the salt tolerance identification at the seedling stage.

[0012] Furthermore, the germination paper is absorbent paper.

[0013] Furthermore, the soybean growth conditions set in the artificial climate chamber include a light intensity of 10,000 lux, a photoperiod of 12 hours of light and 12 hours of darkness, and a temperature of 25°C.

[0014] Furthermore, the matrix is vermiculite.

[0015] The beneficial effect of the present invention is that the method for identifying salt tolerance of soybeans of the present invention realizes a more comprehensive and accurate evaluation of the salt tolerance of soybean varieties by setting up simultaneous identification of salt tolerance of soybean resources at the budding stage and the seedling stage and changing the identification method. It can improve efficiency, save space and consumables when carrying out large-scale identification, classify the salt tolerance of different soybean resources, improve the scientific nature of the evaluation, and can be used to screen breeding materials for soybean salt tolerance breeding. DETAILED DESCRIPTION

[0016] The present invention will be further described in detail below with reference to the embodiments. It should be understood that the specific embodiments described herein are only used to explain the relevant invention, rather than to limit the invention.

[0017] It should be noted that, in the absence of conflict, the embodiments and features of the embodiments in this application can be combined with each other. The present application will be described in detail below with reference to the embodiments.

[0018] The present invention provides a method for identifying salt tolerance of soybeans, comprising step S1 of identifying salt tolerance of soybeans at the budding stage, step S2 of identifying salt tolerance of soybeans at the seedling stage, and step S3 of calculating the salt tolerance grade of soybeans by using a comprehensive evaluation calculation model based on the identification results of salt tolerance of soybeans at the budding stage and the identification results of salt tolerance of soybeans at the seedling stage.

[0019] Wherein, step S1, identification of salt tolerance of soybeans at the sprout stage, comprises:

[0020] S11, fully soaking multiple pieces of germination paper in clean water and 1.2% NaCl solution, generally for 1 to 2 hours, to allow the germination paper to reach a saturated water absorption state;

[0021] S12. Stack two sheets of saturated germination paper and lay them flat on a clean, disinfected table. Evenly place 50 soybeans on top. Cover the soybeans with another sheet of fully soaked germination paper.

[0022] S13. Loosely roll up the three layers of paper with the soybeans, flatten the ends, and place the roll upright in a ziplock bag, leaving a quarter of the seal open for air circulation to allow the soybeans to germinate.

[0023] S14. Using the soybeans germinated in the germination paper soaked in water as a control group, the relative salt damage index of the soybeans germinated in the germination paper soaked in NaCl solution was calculated based on the measured germination rate of the soybeans and the germination rate of the varieties;

[0024] S15. Determine the salt tolerance level of soybean at the budding stage based on the relative salt damage index of soybean.

[0025] As a preferred embodiment, the germination paper is absorbent paper. In this embodiment, the size of the absorbent paper is 40 cm×25 cm.

[0026] To assess the salt tolerance of soybeans during the germination stage, soybeans were wrapped in germination paper and rolled into a roll. The roll was placed vertically to treat the soybeans with salt stress. After sterilization, the germination paper was thoroughly soaked in deionized water (control group) and a 1.2% NaCl solution. Soybean seeds of uniform plumpness and size were selected, disinfected with 75% ethanol, and then washed with deionized water. The seeds were evenly placed on two layers of moistened germination paper. Another piece of germination paper of the same size was then used to cover the soybeans and loosely rolled into a roll. The roll was placed in a ziplock bag and placed vertically in an artificial climate chamber for 7 days.

[0027] In this example, the entire process of evaluating the salt tolerance of soybeans during the sprouting stage was carried out in an artificial climate chamber, and the growth conditions included a light intensity of 10,000 lux, a photoperiod of 12 hours of light and 12 hours of darkness, and a temperature of 25°C.

[0028] The method for grading the salt tolerance of soybeans during the germination stage is to calculate the relative salt damage index PI based on the germination rate and then grade the salt tolerance during the germination stage to determine the level of salt tolerance of soybeans during the germination stage.

[0029] The calculation formula of relative salt damage index is:

[0030] PI = (CK-B) / CK × 100;

[0031] Among them, PI is the relative salt damage index, CK is the germination rate of the control group, and B is the germination rate of the variety.

[0032] The salt tolerance level of soybean during the germination period is 1. Tolerant (relative salt damage index ≤ 20);

[0033] The salt tolerance level of soybean during the budding period is 2, relatively tolerant (relative salt damage index 20-35);

[0034] The salt tolerance level of soybean during the budding period is 3, medium tolerance (relative salt damage index 35-65);

[0035] The salt tolerance level of soybean during the budding period is 4, relatively sensitive (relative salt damage index 65-90);

[0036] The salt tolerance level of soybean during the germination period is 5, sensitive (relative salt damage index>90).

[0037] Step S2, identification of salt tolerance at the seedling stage, includes:

[0038] S21. placing soybeans in the substrate of the cultivation pot;

[0039] S22, placing the cultivation pot in an artificial climate chamber; after the first pair of true leaves of the soybeans in the cultivation pot unfold, watering the soybeans with 0.6% NaCl solution for 14 days;

[0040] S23. Determine the salt tolerance level of soybeans at the seedling stage based on the visual inspection results of the soybean plant growth and the number of damaged leaves.

[0041] As a preferred embodiment, the matrix is vermiculite.

[0042] Salt tolerance of soybean seedlings was assessed using indoor potting. First, small square pots with holes were placed on a tray. Vermiculite substrates of equal mass were placed in the small pots. Randomly selected soybean seeds of equal plumpness were disinfected with 75% ethanol and then rinsed with deionized water. These seeds were placed in small square pots and then covered with 0.5 cm of vermiculite. After the first pair of true leaves of the soybeans unfolded, salt treatment was performed using a 0.6% NaCl solution. An equal amount of clear water was used as a control. Water was replenished every three days based on water loss for 14 days.

[0043] In this embodiment, the salt tolerance of soybeans at the seedling stage was assessed in an artificial climate chamber. Specifically, the soybean growth conditions set in the artificial climate chamber included a light intensity of 10,000 lux, a photoperiod of 12 hours of light and 12 hours of darkness, and a temperature of 25°C.

[0044] The salt tolerance grading method for soybean seedlings is to visually grade based on salt damage symptoms, record the results, and grade the soybeans according to their growth conditions.

[0045] Identification of salt tolerance of soybean at seedling stage: Level 1, tolerant (normal plant growth, green leaves);

[0046] The salt tolerance level of soybean seedlings is 2, relatively tolerant (plant growth is basically normal, 1 / 3 or less of the leaves are affected);

[0047] The salt tolerance level of soybean seedlings is assessed as 3, moderate tolerance (plant growth is inhibited, 1 / 2 or less of the leaves are affected);

[0048] The soybean seedling stage identification salt tolerance level 4, relatively sensitive (plant growth is severely inhibited, 2 / 3 or more of the leaves are affected, or only the upper leaves survive);

[0049] The soybean seedling stage salt tolerance level is 5, sensitive (plant death).

[0050] Step S3 includes calculating the salt tolerance grade of soybeans using a comprehensive evaluation calculation model based on the weights of the soybean salt tolerance identification at the germination stage and the soybean salt tolerance identification at the seedling stage.

[0051] Specifically, the comprehensive evaluation calculation model is:

[0052] N = X × a + Y × b;

[0053] Among them, N is the salt tolerance level of soybean, X is the salt tolerance level of soybean at the germination stage, a is the weight of the salt tolerance identification of soybean at the germination stage, Y is the salt tolerance level of soybean at the seedling stage, and b is the weight of the salt tolerance identification at the seedling stage.

[0054] In this example, the salt tolerance of 12 soybean germplasms at the germination and seedling stages was calculated using the CRITIC weighting method. The result showed that the value of a was 44%; the value of b was 56%.

[0055] The salt tolerance level of soybean is 1. tolerant (N≤1);

[0056] The salt tolerance level of soybean is 2, relatively tolerant (1<N≤2);

[0057] The salt tolerance level of soybean is 3, medium tolerance (2<N≤3);

[0058] The salt tolerance level of soybean is 4, relatively sensitive (3<N≤4);

[0059] The salt tolerance level of soybean is 5, sensitive (N>4).

[0060] The table below shows the results of 12 soybean salt tolerance evaluations. These 12 soybean germplasm samples were bred or collected by agricultural science research institutes in the Jiangsu Yangtze River region between 2022 and 2023. Their salt tolerance was evaluated that year using the test conditions of this patent.

[0061]

[0062]

[0063] The method for identifying salt tolerance of soybeans of the present invention realizes a relatively comprehensive and accurate assessment of the salt tolerance of soybean varieties by setting up simultaneous identification of the salt tolerance of soybean resources at the budding stage and the seedling stage and changing the identification method. It can improve efficiency, save space and consumables when carrying out high-throughput identification, classify the salt tolerance of different soybean resources, improve the scientific nature of the evaluation, and can be used to screen breeding materials for soybean salt tolerance breeding.

[0064] Traditional identification is carried out in culture dishes, but the internal space of the culture dishes is limited. A commonly used 9cm culture dish can only accommodate about 10 soybean seeds. The amount of seeds is small, and if large-scale screening is required, a large amount of consumables and space are required. In addition, the soybean roots in the culture dishes cannot grow normally. After 7 days of growth, the growth state of the seedlings is inconsistent with the natural conditions, the simulation is insufficient, and local salt distribution is prone to unevenness. Compared with traditional culture dish identification, the present invention adopts vertical paper rolls to culture soybean germination, which is highly efficient in salt-tolerant germplasm screening. A single roll of germination paper can hold 50 seeds, greatly improving the screening efficiency. Moreover, since the paper roll is placed vertically, the root system can grow normally, which is consistent with the natural state. The state of each seed can be clearly observed. Therefore, paper roll germination is more efficient in screening salt-tolerant soybeans during the bud stage.

[0065] Furthermore, the present invention's method for identifying soybean salt tolerance at the seedling stage employs a visual grading method for salt tolerance screening. Compared to the traditional, cumbersome procedures of using seed bags and measuring true leaf rate, cotyledon emergence rate, root length, and germination rate, the present invention observes the growth of soybean seedlings and visually grades them based on salt damage symptoms, enabling rapid and efficient screening of salt-tolerant germplasm. This significantly improves identification efficiency, reduces costs, and offers strong applicability, making it highly practical for large-scale screening.

[0066] The above description is merely a preferred embodiment of the present application and an illustration of the technical principles employed. Those skilled in the art should understand that the scope of the invention herein is not limited to the technical solutions formed by the specific combination of the above-mentioned technical features, but also encompasses other technical solutions formed by any combination of the above-mentioned technical features or their equivalents without departing from the inventive concept. For example, a technical solution formed by replacing the above-mentioned features with (but not limited to) technical features having similar functions disclosed in this application.

Claims

1. A method for identifying salt tolerance of soybeans, characterized in that: Including identification of salt tolerance of soybean at germination stage and identification of salt tolerance at seedling stage; The identification of soybean salt tolerance during the germination stage comprises soaking a plurality of germination papers in clean water and a 1.2% NaCl solution, respectively; evenly placing soybeans on two layers of germination paper and covering them with another layer of germination paper; rolling the paper with the soybeans into a roll and vertically placing it in a ziplock bag, maintaining humidity, so that the soybeans germinate; using the soybeans germinated in the germination paper soaked in clean water as a control group, and calculating the relative salt damage index of the soybeans germinated in the germination paper soaked in the NaCl solution based on the measured germination rate and variety germination rate of the soybeans; and determining the salt tolerance level of the soybeans during the germination stage based on the relative salt damage index of the soybeans; The seedling stage salt tolerance assessment includes placing soybeans in a substrate in a cultivation pot; placing the cultivation pot in an artificial climate chamber; watering the soybeans in the cultivation pot with a 0.6% NaCl solution for 14 days after the first pair of true leaves of the soybeans unfold; and determining the seedling stage salt tolerance level of the soybeans based on visual inspection of plant growth and the number of damaged leaves. Based on the weights of the soybean salt tolerance assessment at the germination stage and the soybean salt tolerance assessment at the seedling stage, the soybean salt tolerance grade is calculated using a comprehensive evaluation calculation model, and the comprehensive evaluation calculation model is: N = X × a + Y × b; Among them, N is the salt tolerance grade of the soybean, X is the salt tolerance grade of the soybean at the germination stage, a is the weight of the salt tolerance identification of the soybean at the germination stage, Y is the salt tolerance grade of the soybean at the seedling stage, and b is the weight of the salt tolerance identification at the seedling stage.

2. The method for identifying soybean salt tolerance according to claim 1, wherein The germination paper is absorbent paper.

3. The method for identifying soybean salt tolerance according to claim 1, wherein The soybean growth conditions set in the artificial climate chamber include a light intensity of 10,000 lux, a photoperiod of 12 hours of light and 12 hours of darkness, and a temperature of 25°C.

4. The method for identifying soybean salt tolerance according to claim 1, wherein The matrix is vermiculite.

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