Method for rapidly screening shade-tolerant soybean variety

Abstract

The invention provides a method for rapidly screening shade-tolerant soybean varieties. The invention establishes a method for rapidly identifying and screening shade-tolerant soybean varieties, strains and materials under the conditions of low blue light (LBL) and low red light / far-red light ratio (Low R / FR) in a greenhouse. According to the method for rapidly screening the shade-tolerant soybean variety, the shade tolerance of any soybean variety, strain, material and the like can be rapidly judged, the growth condition of soybeans under soybean close planting or corn-soybean composite planting does not need to be observed on the spot, the identification cost can be saved, and the identification time can also be saved.

Description

Technical Field

[0001] The invention relates to the technical field of plant variety breeding, in particular to a method for rapidly screening shade-tolerant soybean varieties. Background Art

[0002] Soybean is an important food and cash crop in my country, providing not only a high-quality source of protein for humans but also a significant source of feed protein for livestock and other production processes. In recent years, my country's demand for soybeans has continued to rise, necessitating an urgent need to increase soybean yields. Corn-soybean intercropping is one of the primary approaches to increasing soybean yields and is currently being actively promoted in my country. However, soybeans are photoperiod-sensitive plants. Shading from corn can cause soybeans to develop symptoms such as slender stems and reduced branching, known as soybean shade avoidance, which can lead to lodging and reduced yields. Therefore, selecting soybean varieties with strong shade tolerance and minimizing shade avoidance are prerequisites for the successful promotion of corn-soybean intercropping and increased soybean yields, and are therefore crucial for ensuring my country's soybean food security. However, rapid screening techniques for soybean shade tolerance have not yet been established. Summary of the Invention

[0003] The purpose of the present invention is to provide a method for quickly screening shade-tolerant soybean varieties.

[0004] Based on existing research results, the present invention establishes a method for rapidly identifying and screening shade-tolerant soybean varieties, lines, and materials under low blue light (LBL) and low red light / far-red light ratio (Low R / FR) conditions in a greenhouse.

[0005] In order to achieve the purpose of the present invention, the present invention provides a method for quickly screening shade-tolerant soybean varieties, using scheme I and / or scheme II to screen shade-tolerant soybean varieties; Scheme I includes the following steps: (1) Sowing soybeans in pots under ordinary LED lighting conditions. After emergence, plants with inconsistent growth conditions were cut off, and plants with consistent growth conditions such as plant height were retained. (2) 12-14 days after sowing, the plants were divided into treatment groups ① and ②, respectively, and continued to grow for 12-14 days. (3) After 12-14 days of treatment, the plant height or leaf senescence of treatment group ① and treatment group ② were measured respectively. Compared with treatment group ①, the plants in treatment group ② with no significant change in plant height and / or slower leaf senescence were considered to be shade-tolerant plants. Option II includes the following steps: 1) Sow soybeans in pots under ordinary LED lighting conditions. After emergence, cut off plants with inconsistent growth status and retain plants with consistent growth status such as plant height; 2) After the second trifoliate compound leaf of the plant was fully expanded, the plants were divided into two groups: continued treatment under ordinary LED lighting conditions and treatment under LED lights with far-red lamp beads with a low red light / far-red light ratio (i.e., half of the plants continued to be treated under ordinary LED lighting conditions, and the other half were treated with LED lights with far-red lamp beads with a low red light / far-red light ratio), which were recorded as treatment group ③ and treatment group ④ respectively; 3) After the 8-hour darkness period, turn on the LED light for 4-6 hours. Measure the leaf angle of the first trifoliate leaf in both treatment groups ③ and ④. Plants in treatment group ④ that show no significant change in leaf angle compared to treatment group ③ are considered shade-tolerant. The leaf angle measured is the angle between the petiole of the first trifoliate leaf and the main stem.

[0006] Furthermore, in Scheme I, the changes in leaf chlorophyll content in the two treatment groups of plants were used as indicators reflecting leaf senescence.

[0007] Furthermore, in Scheme I, the conditions for ordinary LED lighting (white light treatment) were: 16 h of light and 8 h of darkness, and the total light intensity of the LED light was 400-500 μmol m -2 s -1 , the blue light intensity is 100-150 μmol m -2 s -1 (The total light intensity of the LED lamp is preferably 500 μmolm -2 s -1 , the blue light intensity is 100 μmol m -2 s -1 ).

[0008] Furthermore, weak blue light conditions refer to: adding a yellow filter to the LED light, while keeping the total light intensity unchanged, the blue light intensity is reduced by 20-50 times, that is, the total light intensity of the LED light is 400-500 μmol m -2 s -1 , blue light intensity 3-4μmolm -2 s -1 (Preferably, the total light intensity of the LED lamp is 500 μmol m -2 s -1 , the blue light intensity dropped to 3 μmol m -2 s -1 ). LED light irradiation, 16 hours of light, 8 hours of darkness.

[0009] Furthermore, in Scheme I, the chlorophyll content in a single soybean leaf was measured using a chlorophyll content meter, and the chlorophyll content was statistically calculated using the Student t-test.

[0010] Furthermore, the statistical methods used in determining whether the change in plant height was significant in Scheme I and whether the change in leaf angle was significant in Scheme II were both Student t-test.

[0011] Furthermore, in scheme II, the conditions of treatment group ③ were as follows: ordinary LED lighting conditions: 16 h light, 8 h dark, and the total light intensity of the LED lamp was 400-500 μmol m -2 s -1 , the ratio of red light to far red light is 1-2 (preferably the total light intensity of the LED lamp is 500 μmol m -2 s -1 , the ratio of red light to far-red light is 1.2).

[0012] Furthermore, the conditions of treatment group ④ were as follows: the total light intensity of the LED lamp with far-red light beads was 400-500 μmol m -2 s -1 The ratio of red light to far-red light is 0.1-0.5 (preferably the total light intensity of the LED lamp with far-red light beads is 400 μmol m -2 s -1 , the ratio of red light to far-red light is 0.1). The LED light was used for 16 h of light and 8 h of darkness.

[0013] In the present invention, soybeans are grown in a greenhouse. Preferably, the temperature of the greenhouse is 22-25°C.

[0014] The present invention adopts a potting method to sow soybeans, and the sowing amount of soybeans is 5-10 seeds per pot.

[0015] By means of the above technical solution, the present invention has at least the following advantages and beneficial effects: The method for rapid screening of shade-tolerant soybean varieties provided by the present invention can quickly judge the shade tolerance of any soybean variety, strain, material, etc., without the need to observe the growth conditions of soybeans under dense soybean planting or corn-soybean composite planting in the field, which can save identification costs and identification time. At the same time, the present invention refers to the actual changes in the field spectrum during shading under corn-soybean strip composite planting conditions, determines the spectral changes of shade tolerance identification in the greenhouse, effectively simulates the reduction of blue light and the reduction of red light / far-red light ratio under shading conditions in the field, and increases the practical application value of the present invention. LED lamps are easy to adjust and accurately control light intensity, while saving energy and electricity, and are not prone to high temperatures, which can reduce the temperature control cost of the greenhouse. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 The spectral changes under the conditions of corn-soybean composite planting in the field measured by the present invention are shown.

[0017] Figure 2 This is the spectrum composition after the blue light in the present invention is reduced by 27 times.

[0018] Figure 3 The LED light panel with adjustable light intensity is customized for the present invention.

[0019] Figure 4 This is the LED light board with a yellow filter added in the present invention.

[0020] Figure 5 The present invention is to perform low blue light treatment on a single leaf.

[0021] Figure 6 The LED light source containing far-infrared light is customized for the present invention.

[0022] Figure 7 This is the leaf angle measurement method of the present invention.

[0023] Figure 8 This is a diagram showing the planting of two soybean varieties in a preferred embodiment of the present invention.

[0024] Figure 9 Four soybean varieties in a preferred embodiment of the present invention are shown below LBL.

[0025] Figure 10 These are photos and statistical data of plant height under LBL in a preferred embodiment of the present invention.

[0026] Figure 11 These are photos and statistical data of chlorophyll content under LBL in a preferred embodiment of the present invention.

[0027] Figure 12 This is an LED light board that does not contain FR lamp beads in the preferred embodiment of the present invention.

[0028] Figure 13 This is an LED light board with FR lamp beads in a preferred embodiment of the present invention.

[0029] Figure 14 This is the change in blade angle under low R / FR conditions in a preferred embodiment of the present invention.

[0030] Figure 15 This is a theoretical diagram for selecting plant height, chlorophyll content and leaf angle as indicators in the present invention. DETAILED DESCRIPTION

[0031] The present invention aims to provide a method for rapidly screening shade-tolerant soybean varieties in view of the shortcomings of existing methods for detecting soybean shade-tolerant phenotypes by using filters or adding FR to simulate a shade environment.

[0032] 1. Existing methods simulate shading in the greenhouse and are not correlated with spectral changes in the actual field environment. The present invention measures the light intensity and spectral changes in the field environment to determine the light intensity changes caused by shading under corn-soybean composite planting conditions. Based on the light intensity changes, the total light intensity in the greenhouse and the light intensity after shading are adjusted, thereby improving the accuracy and practicality of the identification data.

[0033] 2. The existing methods for evaluating soybean shade tolerance do not specify the light intensity and specific shading methods after soybean shading. The present invention is based on the molecular theory of spectral changes after soybean shading, and simulates the shading environment by finely regulating the blue light intensity and red light / far-red light ratio.

[0034] 3. Existing methods for evaluating soybean shade tolerance require data collection and statistics after the entire soybean growth period, which takes 3-6 months. The present invention can quickly determine soybean shade tolerance in just 24-28 days.

[0035] 4. Existing methods do not specify specific indicators for shade tolerance screening. Based on existing research results, the present invention determines that plant height and leaf senescence under LBL and leaf angle under Low R / FR are indicators for shade tolerance screening.

[0036] 5. Existing phenotypic identification methods do not clearly describe the experimental process and evaluation methods, such as plant planting time, measurement methods, and spectral composition. This invention will clarify the specific measurement indicators and measurement methods to enhance the operability of screening.

[0037] 6. Existing identification methods do not clearly indicate how to reduce R / FR. The present invention specifies how to increase far-red light to create a low R / FR environment.

[0038] 7. Existing technical methods do not involve the measurement method of leaf angle under low R / FR. The present invention will specifically refine the measurement and statistical method of leaf angle. The present invention adopts the following technical solutions: (1) The present invention first measured the total light intensity in the field, the light intensity between corn and soybean, and the light intensity variation between soybean and soybean under the condition of corn-soybean strip composite planting ( Figure 1 At noon in August 2023, the total light intensity PAR in the fields in Beijing is 1800 μmol m -2 s -1 The blue light intensity is about 480 μmol m -2 s -1 The red light intensity is 700 μmol m -2 s -1 The far-red light intensity is 350 μmol m -2 s-1 , the ratio of red light to far red light is 2 ( Figure 1 A). In corn-soybean strip planting conditions, measurements were taken between corn and soybeans and between soybeans ( Figure 1 D1), and found that the total light intensity between corn and soybean was 70 μmol m -2 s -1 Blue light is only about 10 μmol m -2 s -1 It is about 50 times lower than that in normal sunlight, and the ratio of red light to far-red light is about 0.3 ( Figure 1 B); between soybeans and soybeans ( Figure 1 D2), blue light is only 25 μmol m -2 s -1 It has dropped by nearly 20 times, and the ratio of red light to far red light is 0.5 ( Figure 1 C).

[0039] (2) Since the light intensity of LED lamps in greenhouses is limited, too high light intensity will lead to excessive temperature of the lamp group and greenhouse, increasing the cost of temperature control. In the present invention, the total light intensity of LED lamps in the greenhouse is controlled at 400-500 μmol m -2 s -1 The blue light intensity is controlled at 100-150 μmol m -2 s -1 According to the measurement results in the field, under shade conditions, the blue light intensity is reduced by 20-50 times. In the present invention, the blue light intensity is reduced by 27 times after adding the yellow filter, and is controlled at 3-4 μmol m -2 s -1 , adjust the control total light intensity unchanged ( Figure 2 ).

[0040] (3) According to existing theoretical research results, low blue light and low red light / far-red light ratio are the main light signals for plants to perceive shade under shade conditions. Low blue light signal mainly induces soybean main stem elongation and leaf senescence; low R / FR mainly induces changes in leaf angle ( Figure 15 (Lyu et al., 2021). Therefore, in this study, plants with no significant change in plant height under LBL (Low Blue Light, LBL) and slowed leaf senescence were defined as shade-tolerant plants. Plants with no significant decrease in leaf angle under low R / FR were also defined as shade-tolerant plants.

[0041] (4) Under simulated LBL conditions, the plant height and chlorophyll content were determined as follows: ①Simulate LBL shading signal The custom-made LED light was adjusted to 500 μmol m -2 s -1 ( Figure 3 ), add two layers of yellow filters under the LED light ( Figure 4 ). Increase the voltage to adjust the total light intensity to 500 μmol m -2 s -1 , the blue light intensity dropped to 3 μmol m -2 s -1 ( Figure 4 ). After 12-14 days of treatment, plant height was measured using a ruler and the results were statistically analyzed.

[0042] ②Chlorophyll content determination method Two layers of yellow filters were used to cover a single soybean leaf, specifically filtering out the blue light in white light to simulate the weak blue light conditions under shade conditions ( Figure 5 ); Use two layers of white filter film to cover another single leaf as a control to eliminate the error caused by the influence of light on the leaf after adding two layers of yellow filter. Measure the chlorophyll content after 12-14 days of treatment.

[0043] The chlorophyll content of single soybean leaves was measured using a chlorophyll meter (SPAD-502, Osaka, Japan). The chlorophyll content was statistically analyzed using the Student t-test. Significant differences of P < 0.001 are ***, P < 0.01 are **, P < 0.05 are *, and P > 0.05 is not indicated.

[0044] (5) Simulating low R / FR shade environment Use a customized LED light board and add two rows of FR lamp beads to the light board ( Figure 6 Adjust the total light intensity to 400-500 μmol m -2 s -1 , the R / FR ratio is 1-2. Turn on the FR lamp beads and increase the FR light intensity to reduce the R / FR light ratio to 0.1-0.5, simulating a low R / FR shade environment.

[0045] (6) Period and method of leaf angle measurement After the second trifoliate leaf of the plant to be measured is fully expanded and 4 hours after the light is turned on, use a protractor to measure the leaf angle of the first trifoliate leaf ( Figure 7 ). Measure more than 10 plants and count the leaf angles.

[0046] The following examples are used to illustrate the present invention but are not intended to limit the scope of the present invention. Unless otherwise specified, the technical means used in the examples are conventional means well known to those skilled in the art, and the raw materials used are all commercially available products.

[0047] It should be noted that different soybean varieties were used in Examples 1 and 2. Soybean varieties 1, 2, and 3 in Example 1 are different from varieties 1 and 2 in Example 2. Furthermore, varieties 1, 2, and 3 used in chlorophyll measurements are not the same as varieties 1, 2, and 3 used in plant height measurements.

[0048] Example 1 Determination of shade tolerance by measuring plant height and chlorophyll content under LBL Seven soybean varieties were grown under long-day white light conditions (the total light intensity of LED lamps was 500 μmol m -2 s -1 , the blue light intensity is 100 μmol m -2 s -1 ; Long-day refers to LED light exposure, 16 hours of light, 8 hours of darkness) conditions; one shelf has four black bottom trays, each tray holds two pots (21 cm × 20 cm); 4 pots for each material, 10 seeds / pot ( Figure 8 , showing 2 varieties); 7-10 days after planting, cut off the seedlings with inconsistent growth height, leaving 5 plants per pot; 14 days after planting, transfer the two pots to weak blue light conditions (add two layers of yellow filters under the LED light, adjust the total light intensity by voltage, and reduce the blue light intensity to 3μmolm -2 s -1 ; 16h light, 8h dark) ( Figure 9 , showing 4 varieties). After 14 days, plant height was measured with a ruler and photographed ( Figure 10 ).

[0049] The same planting method as plant height measurement was used. After the seedlings grew for 14 days, two layers of white filters and yellow filters were fixed on two single leaves respectively. After another 14 days of growth, the chlorophyll content was measured and the statistical results were calculated ( Figure 11 ).

[0050] The above results indicate that, among the seven varieties measured for plant height, variety 1 showed significant changes in plant height, indicating a shade-intolerant phenotype. The remaining six varieties showed no significant changes in plant height, indicating good potential for shade tolerance. Among the five varieties measured for chlorophyll content, variety 1 showed no significant changes in chlorophyll content. Varieties 2 and 3 showed accelerated senescence, while varieties 4 and 5 showed slowed senescence. Varieties 4 and 5 demonstrate good potential for shade tolerance.

[0051] Example 2 Measurement of leaf angle at low R / FR The soybean varieties to be tested were planted under normal LED (the total light intensity of LED lamp was 400 μmol m -2 s -1, R / FR ratio is 1.2; 16h light, 8h dark, Figure 12 ) and LED lamps with far-infrared light beads (the total light intensity of LED lamps with far-infrared light beads is 400 μmol m -2 s -1 , R / FR ratio is 0.1; 16h light, 8h dark) ( Figure 13 ), plant 3 plants in each pot. After the second trifoliate leaf of the plant is fully expanded, 4 hours after turning on the light, use a protractor to measure the leaf angle of the first trifoliate leaf and take photos of the plants (take 1 out of 3 plants to take a photo), and count the changes in leaf angle ( Figure 14 ). The statistical results show that the leaf angle of test variety 1 changed significantly (P<0.001), and the leaf angle of test variety 2 changed more significantly (P<0.01) (To determine the shade tolerance of a variety by leaf angle, it is necessary to use t-test to judge the significance of the change in leaf angle before and after low R / FR treatment, rather than using the range of leaf angle change. We set P>0.001 as the criterion for determining shade tolerance potential). The degree of change was 36.13° and 9.36°, respectively. Test variety 2 is a variety insensitive to low R / FR and has good shade tolerance potential.

[0052] Although the present invention has been described in detail above using general descriptions and specific embodiments, it will be apparent to those skilled in the art that modifications and improvements may be made based on the present invention. Therefore, such modifications and improvements, which do not depart from the spirit of the present invention, are intended to be within the scope of protection claimed herein.

Claims

1. A method for rapidly screening shade-tolerant soybean varieties, characterized in that: Using Scheme I and / or Scheme II to select shade-tolerant soybean varieties; Scheme I includes the following steps: (1) Sowing soybeans in pots under ordinary LED lighting conditions. After emergence, plants with inconsistent growth status were cut off, and plants with consistent height growth status were retained. (2) 12-14 days after sowing, the plants were divided into two groups: treatment group ① and treatment group ②, which were treated under normal LED lighting conditions and weak blue light conditions, respectively, and continued to grow for 12-14 days; (3) After 12-14 days of treatment, the plant height or leaf senescence of treatment group ① and treatment group ② were measured respectively. Compared with treatment group ①, the plants in treatment group ② with no significant change in plant height and / or slower leaf senescence were considered to be shade-tolerant plants. Option II includes the following steps: 1) Sow soybeans in pots under ordinary LED lighting conditions. After emergence, cut off plants with inconsistent growth status and retain plants with consistent height growth status; 2) After the second trifoliate leaf of the plant fully expanded, the plants were divided into two groups: continued treatment under normal LED lighting conditions and treatment under LED lighting with a low red light / far-red light ratio and far-red light beads, respectively recorded as treatment group ③ and treatment group ④; 3) After the LED light was turned on for 4-6 hours, the leaf angles of the first trifoliate leaves of the plants in treatment groups ③ and ④ were measured respectively. Compared with treatment group ③, the plants in treatment group ④ with no significant change in leaf angle were the plants with strong shade tolerance.

2. The method according to claim 1, characterized in that In scheme I, the changes in leaf chlorophyll content in the two treatment groups were used as indicators reflecting leaf senescence.

3. The method according to claim 1, characterized in that In scheme I, the normal LED lighting conditions are: 16 h light, 8 h dark, and the total light intensity of the LED lamp is 400-500 μmol m -2 s -1 , the blue light intensity is 100-150 μmol m -2 s -1 .

4. The method according to claim 3, characterized in that Weak blue light conditions refer to adding a yellow filter to the LED light, with a total light intensity of 400-500 μmol m -2 s -1 , the blue light intensity is reduced by 20-50 times; 16h light, 8h dark.

5. The method according to claim 4, characterized in that The blue light intensity was reduced to 3-4 μmol m -2 s -1 .

6. The method according to claim 1, characterized in that In scheme I, the chlorophyll content of a single soybean leaf is measured using a chlorophyll content meter, and the chlorophyll content is statistically calculated using the Student t-test; and / or, The statistical methods used to determine whether the change in plant height was significant in Scheme I and whether the change in leaf angle was significant in Scheme II were both Student t-test.

7. The method according to claim 1, characterized in that In Scheme II, the normal LED lighting conditions are: 16 h light, 8 h dark, and the total light intensity of the LED lamp is 400-500 μmol m -2 s -1 , the ratio of red light to far-red light is 1-2.

8. The method according to claim 7, characterized in that The conditions of treatment group ④ were as follows: the total light intensity of the LED lamp with far-red light beads was 400-500 μmol m -2 s -1 , the ratio of red light / far-red light is 0.1-0.5; 16h light, 8h dark.

9. The method according to any one of claims 1 to 8, characterized in that Soybeans are grown in greenhouses; Preferably, the temperature of the greenhouse is 22-25°C.

10. The method according to any one of claims 1 to 8, characterized in that Soybeans are sown in pots, with a sowing rate of 5-10 seeds per pot.

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