Rice seed vigor detection method based on oxygen sensing

Abstract

The invention provides a rice seed vigor detection method based on oxygen sensing, and the rice seed vigor detection method based on oxygen sensing comprises the steps of pretreatment, detection and statistics, and specifically comprises the following steps: selecting full, consistent, insect-hole-free and mildew-free rice seeds, and carrying out aging treatment to obtain pre-detected rice seeds a; agar is added into distilled water to prepare an agar solution b with the concentration of 0.4-0.6%; adding 700-900L of the agar solution b into a sample tube, adding a single seed of the to-be-detected rice seed a into the sample tube after solidification, transferring into a seed vitality instrument, monitoring an oxygen consumption curve in real time, and screening out two parameters, namely an initial metabolic rate (SMR) and an oxygen metabolic rate (OMR); the SMR and OMR indexes are used for judging the seed vigor.

Description

Technical Field

[0001] The present invention relates to the field of detection technology, and in particular to a rice seed vitality detection method based on oxygen sensing. Background Art

[0002] Seed vigor refers to the potential for seeds to germinate under actual field conditions and its correlation with yield and related agronomic indicators. It is not only closely linked to crop production but also to physiological and biochemical processes such as seed development, maturation, germination, storage life, and deterioration. As a key assessment criterion for seed quality, seed vigor plays a central role in seed quality assessment and seed trading. Previously, one of the most commonly used methods for testing seed vigor was the seed germination test. However, traditional germination tests are time-consuming and labor-intensive, making them difficult to meet the current demand for rapid, efficient, and accurate seed viability assessment in modern agricultural production. Therefore, the development of a product that can address these technical issues is crucial. Summary of the Invention

[0003] The purpose of the present invention is to provide a rice seed vitality detection method based on oxygen sensing.

[0004] The object of the present invention is achieved in this way. The wheat vitality detection method based on oxygen sensing includes pre-processing, detection and statistical steps, specifically including: A. Pre-treatment: Select plump, uniform, insect-free, and mildew-free rice seeds and perform aging treatment to obtain pre-test rice seeds a; B. Testing: 1) Add agar to distilled water to prepare agar solution b with a concentration of 0.4-0.6%; 2) Add 700-900 µL of agar solution b to a sample tube. After solidification, add a single seed from pre-tested rice seed a to the sample tube. Transfer the sample to a seed viability meter to monitor the oxygen consumption curve in real time, screening out the initial metabolic rate (SMR) and oxygen metabolic rate (OMR). C. Statistics: SMR and OMR indicators are used to determine seed vitality.

[0005] This invention constructs an innovative system based on oxygen sensing technology for rice seed viability detection. Its core technical features include: using a proprietary combination of 0.5% agar medium and 800μl volume (adapted to the physiological characteristics of rice to ensure 100% suspension of seeds on the agar surface), monitoring the oxygen consumption curve at a constant temperature of 25°C using a seed viability meter, and screening the initial metabolic rate (SMR) and oxygen metabolic rate (OMR) as key judgment indicators (excluding fluctuating parameters such as IMT / R50 / AUC50 / RGT). Combined with a validation model of gradient aging for 7 / 14 / 30 days at 40°C and 75% humidity, a quantitative correlation between SMR / OMR values ​​and viability level is established (for example, an OMR ≤ 0.004 indicates loss of viability). The technology's benefits include: breaking through testing bottlenecks—replacing traditional germination tests that require more than seven days, enabling automated high-throughput testing within seven days (increasing throughput by 40-fold and reducing labor costs by 90%), and addressing germination anomalies caused by seed sinking to the bottom in low-concentration agar. It also improves accuracy and reliability, achieving a SMR / OMR parameter error of <2% (a 2.5-fold improvement compared to the ±5% accuracy of germination rate), and ensuring early warning of vigor deterioration through aging verification (detecting a significant downward trend after just seven days of aging). It also forms a closed-loop decision-making process, integrating the entire chain of "media formulation-equipment parameters-algorithms-verification standards" to output vigor levels that directly guide seed bank conservation strategies (e.g., "OMR < 0.01 → prioritize propagation"), providing efficient technical support for the safe preservation of germplasm resources. This system is the first to achieve crop-specific adaptation (media volume / parameter set differs from wheat), and demonstrates the universal methodology of "media volume matching seed size" and "parameter validity verification through aging," providing a standardized paradigm for the development of vigor detection systems for other crops.

[0006] (I) Testing medium system, specific concentration and volume combinations: Rice seeds (Lianghe Mangdong Huangkenuo) require a 0.5% agar concentration + 800μl volume as the germination medium. Technical results: The oxygen consumption curve exhibits a stable "inverted S-shaped" shape, significantly superior to other combinations (such as 0.1% / 0.3% concentration or 400 / 1000μl volume). The seeds are completely placed on the agar surface, avoiding sinking or suspension, ensuring a consistent germination environment.

[0007] (II) Key testing indicators: Core parameters: Initial metabolic rate (SMR) and oxygen metabolic rate (OMR) are core indicators for determining rice vitality. Excluded parameters: IMT, R50, AUC50, and RGT were excluded as valid indicators due to large fluctuations in aging experiments or failure to conform to the law of vitality changes.

[0008] (III) Supporting Verification Methods: Aging treatment: 7, 14, and 30 days at 40°C and 75% humidity as a standard for verifying vigor changes. Data Correlation: Decreasing trends in SMR and OMR values ​​are significantly correlated with decreased germination potential and rate.

[0009] Advantages of the present invention: 1. Addressing the shortcomings of traditional technologies and replacing time-consuming germination experiments: Oxygen sensing technology (Q2) enables high-throughput testing within 7 days, overcoming the traditional methods' high labor reliance and requiring more than 7 days. Improved accuracy: Under the conditions of 0.5% agar + 800μl, the SMR / OMR indicators showed a stable response to aging, with a low false positive rate (Table 3-6).

[0010] 2. Crop-specific adaptation, rice-specific system: The medium volume (800μl) differs from that of wheat (1000μl); the parameters (SMR / OMR only) differ from the multi-parameter system (SMR / OMR / R50 / AUC50 / RGT) for wheat. Technical Effect: Optimized for the physiological characteristics of glutinous rice (such as aerobic germination patterns), universal applicability requires further expansion to other varieties.

[0011] 3. Technological integration and innovation: a closed-loop "medium-equipment-algorithm" system: medium formula (0.5% agar) → equipment setting (25°C constant temperature) → parameter extraction (SMR / OMR algorithm) → verification standard (aging gradient); this system standardizes "test conditions-result determination" and provides a reusable technical solution for seed bank vitality monitoring. BRIEF DESCRIPTION OF THE DRAWINGS

[0012] Figure 1 This is a schematic diagram of the oxygen consumption curve of Lianghe Mangdong Huangkenuo in Example 1 of the present invention; Figure 2 This is a schematic diagram of the seed growth in Example 1 of the present invention; Figure 3 This is a bar chart of various indicators of Lianghe Mangdong Huangkeno in Example 1 of the present invention. DETAILED DESCRIPTION

[0013] The present invention is further described below with reference to the embodiments, but the present invention is not limited in any way. Any changes or substitutions made based on the teachings of the present invention fall within the scope of protection of the present invention.

[0014] The wheat vitality detection method based on oxygen sensing of the present invention includes pre-processing, detection and statistical steps, specifically including: A. Pre-treatment: Select plump, uniform, insect-free, and mildew-free rice seeds and perform aging treatment to obtain pre-test rice seeds a; B. Testing: 1) Add agar to distilled water to prepare agar solution b with a concentration of 0.4-0.6%; 2) Add 700-900 μL of agar solution b to the sample tube, and after solidification, add the single seeds of the pre-tested rice seeds a to the sample tube, and transfer to the seed vigor instrument to monitor the oxygen consumption curve in real time, and screen out the initial metabolic rate SMR and oxygen metabolic rate OMR parameters; C. Statistics: using SMR and OMR as indicators for determining seed vigor.

[0015] The aging treatment in step A is to place the rice seeds in a temperature of 30-50℃ and a humidity of 70-80% for aging treatment.

[0016] The aging treatment is to place the rice seeds in a temperature of 35-45℃ and a humidity of 74-76% for aging treatment.

[0017] The aging treatment time is 7-30 days.

[0018] The temperature for real-time monitoring of the oxygen consumption curve in step B2) is 20-30℃.

[0019] The application will be further described below with specific examples: Example 1

[0020] 1. Rice seed vigor analysis 1.1 Oxygen consumption detection analysis of rice seed vigor instrument without aging treatment Figure 1 The oxygen consumption curves of Lianghe Mangdonghuangniao under different germination concentrations and volumes are as follows: Figure 1 a, b, c, and d are the oxygen consumption curves of agar with different volumes and a concentration of 0.1%, Figure 1 e, f, g, and h are the oxygen consumption curves of agar with different volumes and a concentration of 0.3%, Figure 1 i, j, k, and l are the oxygen consumption curves of agar with different volumes and a concentration of 0.5%.

[0021] It is observed that the oxygen consumption curves with a concentration of 0.1% do not well reflect the reverse "S" shape. In addition, under the same concentration conditions, more and more oxygen consumption curves present the reverse "S" shape with the increase of volume. Under the same volume conditions, more and more oxygen consumption curves present the reverse "S" shape with the increase of concentration. Figure 1The i, j, k, and l values ​​generally exhibited a relatively good inverse "S" curve, indicating that an agar concentration of 0.5% and volumes of 400, 600, 800, and 1000 μl were the most suitable conditions for seed viability instrument testing. Based on the oxygen consumption curve, the optimal agar concentration of 0.5% was screened. Further comparison of oxygen consumption indicators at different volumes revealed that there were no significant differences in OMR, R50, AUC50, and RGT values ​​at volumes of 400, 600, 800, and 1000 μl. Furthermore, at 800 μl, the R50, AUC50, and RGT values ​​were all lower than those at the other three volumes. There was a significant difference in SMR values ​​between 400 and 600 μl, but not between 800 and 1000 μl. There was also a significant difference in IMT values ​​between 1000 μl and the other three volumes. In summary, 800 μl at 0.5% was the most suitable volume for seed viability instrument testing.

[0022] Table 1 Parameters for detecting vitality of Huangke glutinous rice in Mangdong, Lianghe Note: The difference is significant at the p = 0.05 level. At the same time, combined with the germination performance of seeds in different media, it can be seen that ( Figure 2 ), in 0.1% agar concentration, most of the seeds of Lianghe Mangdong Huangke glutinous rice have sunk to the bottom of the tube, and the growth potential is uneven; in 0.3% agar concentration, some seeds are still suspended in the agar, of which 1000μl has the best growth potential; in 0.5% agar concentration, the seeds are completely placed on the agar and will not be embedded in the agar. The oxygen consumption curve of 800μl (Table 1, Figure 2 ) and their growth potential results were consistent. This indicates that 0.5% agar (800 μl) is suitable for seed viability testing.

[0023] 1.2 Analysis of oxygen consumption by vigor meter after aging treatment of rice seeds Figure 3 Figure 1 shows the statistical results of different oxygen consumption indicators after aging treatment. Generally speaking, the stronger the seed vigor, the larger the SMR and OMR values, and the smaller the IMT, R50, AUC50, and RGT values. With increasing aging time, seed vigor decreases, with SMR and OMR values ​​showing a downward trend, while IMT, R50, AUC50, and RGT values ​​show an upward trend. Overall, the trends in oxygen consumption indicators after aging as measured by the seed vigor meter are consistent with the observed decrease in vigor of rice seeds after aging.

[0024] A specific comparison of the changes in different oxygen consumption indicators during the aging stage revealed that ( Figure 3 ): When the germination medium concentration was 0.1%, the oxygen consumption indicators SMR, IMT, OMR, R50, AUC50, and RGT showed an overall trend consistent with declining seed viability at different volumes. However, the values ​​fluctuated at specific time points, such as the SMR (0.1%, 800μl). When the germination medium concentration was 0.3%, the oxygen consumption indicators also fluctuated at specific time points at different volumes, such as the OMR (0.3%, various volumes). When the germination medium concentration was 0.5%, the SMR values ​​at the 800μl volume showed a consistent trend, with no fluctuations. This suggests that the 0.5% and 800μl volumes are more suitable for seed viability testing.

[0025] Table 2 Vitality parameter values ​​of Lianghe Mangdong Huangke glutinous rice after different aging treatments in germination medium with an agar concentration of 0.5% and a volume of 800 μl Note: The difference is significant at the p = 0.05 level. Table 2 shows that the SMR values ​​showed significant differences between 0, 7, and 14 days of aging, but not between 14 and 30 days. The SMR values ​​gradually decreased with increasing aging time, consistent with the trend that lower seed vigor results in lower SMR values. Although the IMT values ​​generally increased, they showed a downward trend after 7 days of aging, which is inconsistent with the regular pattern of seed vigor changes. The OMR values ​​showed no significant differences between 0, 7, and 14 days of aging, but showed significant differences between 30 days of aging and 0, 7, and 14 days of aging. Although the R50, AUC50, and RGT values ​​generally increased, they were lower after 7 days of aging than after 0 days, which is inconsistent with the trend that lower vigor results in higher values. Therefore, the SMR and OMR are the most suitable indicators for detecting Huangke Nuo in Mangdong, Lianghe.

[0026] 1.3 Rice seed standard germination test analysis Table 3 shows the statistical results of germination rate and germination potential of rice seeds after different aging treatments. The results show that the germination potential of rice seeds increases with the extension of aging time, and its germination potential drops to 0 after 30 days of aging. The germination rate is also affected by the aging time. With the increase of aging time, the germination rate decreases significantly.

[0027] Comparing the oxygen consumption index test results of the vitality meter with those of the standard germination experiment, it was found that the two test results were consistent, which shows that the seed vitality meter can be used in rice seed vitality detection, and like the standard germination experiment, it has reliable and accurate test results.

[0028] Table 3. Germination potential (rate) of Huangke glutinous rice in Mangdong, Lianghe As can be seen from Table 4, the germination potential of Lianghe Mangdong Huangke glutinous rice gradually decreases with the increase of aging time. There is no significant difference between aging for 0 days and aging for 7 days, but there are significant differences between aging for 0 days and 7 days and aging for 14 days and 30 days; there are significant differences in germination rate at all time points.

[0029] Table 4 Variance analysis of germination potential (rate) of Huangke glutinous rice in Mangdong, Lianghe Note: The difference is significant at the p = 0.05 level.

Claims

1. A rice seed vitality detection method based on oxygen sensing, characterized in that: The wheat vitality detection method based on oxygen sensing includes pre-processing, detection and statistical steps, specifically including: A. Pre-treatment: Select plump, uniform, insect-free, and mildew-free rice seeds and perform aging treatment to obtain pre-test rice seeds a; B. Testing: 1) Add agar to distilled water to prepare agar solution b with a concentration of 0.4-0.6%; 2) Add 700-900 µL of agar solution b to a sample tube. After solidification, add a single seed from pre-tested rice seed a to the sample tube. Transfer the sample to a seed viability meter to monitor the oxygen consumption curve in real time, screening out the initial metabolic rate (SMR) and oxygen metabolic rate (OMR). C. Statistics: SMR and OMR indicators are used to determine seed vitality.

2. The rice seed vitality detection method based on oxygen sensing according to claim 1, characterized in that: The aging treatment described in step A is to place the rice seeds under the conditions of a temperature of 30-50° C. and a humidity of 70-80% for aging treatment.

3. The rice seed vitality detection method based on oxygen sensing according to claim 2, characterized in that: The aging treatment is to place the rice seeds at a temperature of 35-45° C. and a humidity of 74-76% for aging treatment.

4. The rice seed vitality detection method based on oxygen sensing according to claim 2 or 3, characterized in that: The aging treatment time is 7 to 30 days.

5. The method for detecting rice seed vitality based on oxygen sensing according to claim 1, characterized in that: The temperature for real-time monitoring of the oxygen consumption curve in step B2) is 20-30°C.

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