Method for identifying plutella xylostella-resistant germplasm resources of cruciferous vegetable crops

Abstract

The invention discloses a method for identifying plutella xylostella-resistant germplasm resources of cruciferous vegetable crops. The identification method comprises an identification device and an identification method, the identification device is provided with a breathable net on a cover body of a culture dish, agarose, filter paper and moisturizing cotton are combined to construct a proper feeding environment for the plutella xylostella, and a field feeding process is truly simulated under indoor conditions. On the basis of the device, an insect resistance identification method is established, three-instar larvae of plutella xylostella are inoculated and cultured regularly, and larva death number, weight change and leaf damage data are obtained; and further constructing a germplasm resource identification method for resisting the plutella xylostella of the cruciferous vegetable crops, and carrying out quantitative analysis and grade judgment on the insect resistance of the cruciferous vegetable crops by taking the larva death rate, the body weight inhibition rate and the leaf damage rate as evaluation indexes. The identification device and method cooperate with each other to form a standardized cruciferous vegetable crop insect-resistant resource identification system.

Description

Technical Field

[0001] This invention relates to the field of identification technology for insect-resistant resources in agricultural vegetables, and in particular to a method for identifying the insect resistance of cruciferous vegetable crops. Background Technology

[0002] The Brassicaceae family is widely distributed throughout the world, comprising approximately 375 genera and 3,200 species. Among them, those widely cultivated as vegetables are mainly divided into two genera: *Brassica* and *Raphanus* L. *Brassica* vegetables are further divided into three categories: *Brassica oleracea*, *Brassica rapa*, and *Brassica juncea*. *Brassica oleracea* vegetables mainly include cauliflower, broccoli, head cabbage, and kale; *Brassica rapa* vegetables mainly include Chinese cabbage, non-heading Chinese cabbage, and turnips; *Brassica juncea* vegetables mainly include leaf mustard, root mustard, stem mustard, and flowering mustard. The *Raphanus* genus mainly includes wild and cultivated radishes. In today's diet, cruciferous vegetables are receiving increasing attention due to their rich nutritional content and health benefits, and are an indispensable part of daily meals. Cruciferous vegetables can be eaten as vegetables, as well as used as oil and feed. Therefore, they are widely planted in my country and have extremely important economic value.

[0003] The diamondback moth (Plutella xylostella (L.)) belongs to the family Plutellidae in the order Lepidoptera. It is a global agricultural pest that damages cruciferous vegetables, with over 40 host plant species. First-instar larvae feed and burrow into the leaves of host plants, continuously consuming the leaf mesophyll. As they enter the second instar, their appetite increases, leaving only remnants of the leaf epidermis, a phenomenon known as "opening windows." The third and fourth instar larvae are in their voracious feeding phase, with a significantly increased appetite, creating large areas of holes or notches in the leaves. In severe cases, they can even eat the veins, causing a decline in the quality of the host plant and resulting in significant economic losses. It is estimated that the global economic losses caused by diamondback moth damage reach US$4-5 billion annually, while in my country, the annual economic losses are as high as US$770 million.

[0004] Control methods for the diamondback moth can be categorized into agricultural, biological, physical, and chemical control, but chemical control is the primary method in agricultural production. However, the excessive and inappropriate use of chemical pesticides has not only caused environmental pollution and varying degrees of damage to ecosystems but has also led to diamondback moth resistance to almost all pesticides, seriously affecting the control of the moth and food safety. Currently, based on the differences in insect resistance exhibited by different vegetable genotypes, breeding new insect-resistant vegetable varieties is the most economical and effective measure for controlling the diamondback moth. However, it is still unclear which vegetable varieties possess insect resistance among the many available; therefore, the evaluation and identification of vegetable insect resistance is the prerequisite and foundation for insect-resistant vegetable breeding.

[0005] Currently, methods for identifying plant insect resistance include field natural identification, field inoculation with increased insect population pressure, and net-house identification. However, these methods are commonly used in field crops (rice, corn, and wheat), and are time-consuming, labor-intensive, and have long screening cycles. Furthermore, research on insect-resistant breeding in vegetables started relatively late, and insect resistance identification equipment and technologies are scarce, severely hindering the development of vegetable insect-resistant breeding. At the same time, traditional crop insect resistance identification methods are not perfectly suited for vegetable insect resistance identification, suffering from problems such as difficulty in precisely controlling experimental conditions and unscientific identification methods. They lack accurate resistance indicators and parameters, making it impossible to comprehensively and accurately assess the resistance of cruciferous vegetables to the diamondback moth. Therefore, developing an identification method for insect-resistant germplasm resources in cruciferous vegetables is of great significance for the development of vegetable insect-resistant breeding. Summary of the Invention

[0006] The purpose of this invention is to overcome the problems of insufficient equipment, cumbersome operation, time-consuming and laborious statistical analysis, and poor accuracy of evaluation results in the identification of insect-resistant germplasm resources of cruciferous vegetables in existing technologies. This invention provides an device and method for identifying insect-resistant germplasm resources of cruciferous vegetables. The device and method are simple to operate, low in cost, and can accurately reflect the resistance level of cruciferous vegetables to the diamondback moth, screening out susceptible and highly resistant germplasm materials of cruciferous vegetables to the diamondback moth, thus improving the efficiency of insect-resistant germplasm resource identification.

[0007] To achieve the above objectives, the present invention is implemented through the following technical solution:

[0008] In a first aspect, the present invention provides an identification device for insect-resistant germplasm resources of cruciferous vegetable crops, comprising a breathable net, a lid, a 2% agarose matrix, moistened filter paper, a moistening cotton, and a petri dish; the lid is provided with a breathable net; the bottom of the petri dish is covered with a 2% agarose matrix, and moistened filter paper is placed on top of the agarose; the petri dish, the moistened filter paper, the breathable net, and the lid form a sealed space, and a third-instar larva of the diamondback moth and a leaf are placed in the petri dish; the petiole of the leaf is wrapped with a moistening cotton.

[0009] Preferably, the mesh size of the breathable mesh is not less than 100 mesh.

[0010] Preferably, the moisturizing cotton is moistened with sterile water.

[0011] Secondly, the present invention provides a method for identifying insect-resistant germplasm resources of cruciferous vegetable crops, the method being as follows:

[0012] Step S1: Select cruciferous vegetable materials to be identified;

[0013] Step S2: Inoculate the third instar larvae of the diamondback moth onto the leaves of the cruciferous vegetables;

[0014] Step S3: Cultivate the inoculated diamondback moth larvae;

[0015] Step S4: Observe and record the survival and development status of diamondback moth larvae;

[0016] Step S5: Conduct a comprehensive evaluation and data analysis of the diamondback moth resistance of cruciferous vegetable crop materials.

[0017] Preferably, the selection of cruciferous vegetable materials to be identified in step S1 is carried out as follows: different cruciferous vegetable seeds are sown in culture pots, with 3 seeds sown in each culture pot. Then, they are placed in a greenhouse. When the cruciferous vegetables grow to 4 leaves, the weak seedlings are removed and the healthy and strong seedlings are retained. Then, when the seedlings grow to 8 leaves, the 4th-5th leaves are used for insect resistance identification. Each variety is repeated 3 times.

[0018] Preferably, the preparation of the evaluation and identification device described in step S2 is as follows: Select a 90 mm plastic petri dish, place 2% agarose at the bottom of the petri dish, place a filter paper with a diameter of 86 mm on top of the agarose, use a brush to dip in sterile water to moisten the filter paper, cut out the area of ​​40-60 mm in the center of the petri dish lid, and use hot melt glue to stick a 100-mesh breathable mesh to the empty area.

[0019] Preferably, the inoculation of the third instar larvae of the diamondback moth in step S3 is carried out as follows: select third instar larvae of the diamondback moth, inoculate 10 third instar larvae of the diamondback moth into each petri dish, cover the top of the petri dish with absorbent paper, and then cover the petri dish with a lid.

[0020] Preferably, the culture conditions after inoculation of the third instar larvae of the diamondback moth in step S3 are: an artificial climate insect rearing room with an ambient temperature of 25 ± 1℃ and a relative humidity of 65%.

[0021] Preferably, the observation of the survival and development of diamondback moth larvae in step S4 is carried out in the following steps: 24 h, 48 h and 72 h after completing step S3, the diamondback moth larvae in the petri dish are observed, and the number of dead larvae, the weight changes of surviving larvae and the damaged area of ​​cruciferous vegetable leaves are recorded.

[0022] Preferably, the data analysis of cruciferous vegetables' resistance to diamondback moth in step S5 includes the following steps: Based on the number of dead diamondback moth larvae, changes in the weight of surviving larvae, and the area of ​​leaf damage recorded in step S4, the corrected mortality rate of larvae, the rate of change in the weight of surviving larvae, and the leaf damage rate are calculated respectively; then, the above indicators are weighted according to a preset weight ratio to obtain the comprehensive insect resistance index I of the cruciferous vegetable materials. The comprehensive insect resistance index is positively correlated with the mortality and weight changes of larvae and negatively correlated with the degree of leaf damage; finally, based on the numerical range of the comprehensive insect resistance index I, the cruciferous vegetable materials are divided into five levels: highly resistant, resistant, moderately resistant, susceptible, and highly susceptible, thereby achieving a comprehensive evaluation and identification of the resistance of cruciferous vegetables to diamondback moth.

[0023] Thirdly, the insect resistance of cruciferous vegetable varieties is comprehensively evaluated by combining multiple indicators (mortality rate, weight change, and leaf damage). Unlike traditional single-indicator determinations, the comprehensive insect resistance evaluation method of this invention not only improves the accuracy and stability of the evaluation but also makes the results more scientific and reliable.

[0024] Preferably, the calculation of the corrected mortality rate in step S5 is important because insect mortality rate is often used as a basic indicator in crop insect resistance evaluation. However, assessing insect resistance solely based on mortality rate has significant errors because insect mortality rate can be affected by various factors during the experiment. For example, environmental temperature, humidity, and insect population density can all influence mortality rate. Therefore, absolute mortality rate alone cannot accurately reflect plant resistance.

[0025] Corrected mortality rates, by comparing the mortality rates of the experimental and control groups, effectively eliminate the influence of external environmental factors and operational errors, ensuring that the measured mortality rates accurately reflect differences in crop insect resistance. Especially under indoor experimental conditions, even small environmental fluctuations can significantly impact insect survival; corrected mortality rates better control these variables and reduce bias. Calculating corrected mortality rates not only eliminates experimental errors but is also a standard research method to ensure the reliability of results. This method allows researchers to more accurately compare the insect resistance of different crop varieties and ensures high reproducibility of experimental data.

[0026] Preferably, the calculation of the weight change rate described in step S5 provides researchers with information about crop insect resistance mechanisms. For example, some crops may slow insect weight gain by inhibiting insect metabolism, and these inhibitory effects may not immediately lead to death. Therefore, by observing weight changes, researchers can understand how crop resistance substances affect insect physiological processes through different mechanisms, and further analyze their insect resistance mechanisms. The weight change rate, as an indicator of insect health, compensates for the inadequacy of mortality alone. Even if insects survive, their growth and development may be inhibited by crop resistance. The weight change rate quantifies the inhibitory effect of crops on insect growth, rather than just the life or death of insects, making it an indispensable core indicator in insect resistance research.

[0027] Preferably, the calculation of leaf damage rate in step S5 is important because leaves are the main organs for photosynthesis in plants, and insect feeding on leaves directly affects plant growth and yield. Leaf damage rate, as a direct indicator of the extent of insect damage, can accurately assess crop resistance to insects. By measuring the area of ​​damaged leaves, researchers can clearly understand the actual degree of damage caused by insects to crops. Leaf damage rate is not only an external representation of crop insect resistance but also reflects the effectiveness of crop defense responses. Crops reduce insect damage by secreting toxic substances, strengthening physical barriers, or altering metabolic pathways, and leaf damage rate is a direct indicator of the effectiveness of these defense mechanisms. Combining leaf damage rate with mortality rate and weight change rate provides a more comprehensive assessment of insect resistance. Mortality rate reflects insect survival, weight change rate reflects insect growth, and leaf damage rate directly characterizes the extent of insect damage to crops. The combined effect of these three factors allows for a more scientific and accurate assessment of plant insect resistance. It is a standardized method for detecting crop insect resistance responses, helping researchers screen insect-resistant germplasm resources and conduct in-depth research on crop defense mechanisms.

[0028] The present invention has the following beneficial effects:

[0029] (1) The present invention provides an identification device for insect-resistant germplasm resources of cruciferous vegetable crops. The device is made of readily available materials, is easy to assemble, easy to operate, and has low cost. It is suitable for widespread use in laboratories and breeding conditions.

[0030] (2) The present invention provides an identification device for insect-resistant germplasm resources of cruciferous vegetable crops. The device provides 2% agarose that can continuously retain moisture, which changes the defect of short moisturizing time of single-layer filter paper in the past.

[0031] (3) The present invention provides an identification device for insect-resistant germplasm resources of cruciferous vegetable crops. The device has a 40-60 mm area cut out in the center of the lid and a 100-mesh breathable net is attached to it, which increases the breathability of the device and changes the defect that the detection data was inaccurate in the past due to the large number of insects dying due to excessive humidity.

[0032] (4) The present invention provides an identification device for insect-resistant germplasm resources of cruciferous vegetable crops. The petri dish of the device is 90 mm in diameter, which can hold the whole leaf to be tested, thus avoiding the defect of inaccurate test data caused by severe water loss and yellowing of leaves due to leaf cutting in the past.

[0033] (5) This invention uses the device to identify insect-resistant germplasm resources of cruciferous vegetable crops. This method breaks the long-standing dominance of European and American countries in the identification of insect-resistant germplasm resources of crops and creatively provides China's own identification method for insect-resistant vegetable germplasm resources.

[0034] (6) The present invention uses this device to identify insect-resistant germplasm resources of cruciferous vegetable crops. The method selects the third-instar larvae of the diamondback moth, a specialized pest of cruciferous vegetables, as the object. This instar larvae are in the voracious feeding period of field pests and can represent the period when field pests cause the most serious damage, and have obvious representativeness.

[0035] (7) The present invention adopts the method of simultaneously recording the number of dead diamondback moth larvae, changes in body weight, and damaged leaf area, providing more comprehensive and multi-dimensional data support for subsequent insect resistance evaluation.

[0036] (8) This invention introduces a multi-indicator calculation method, such as corrected mortality rate, weight change rate and leaf damage rate, and constructs a comprehensive insect resistance index I through weighting, so that resistance evaluation no longer depends on a single indicator, and the evaluation results are more stable, scientific and quantifiable.

[0037] (9) Based on the comprehensive insect resistance index, five levels are divided into high resistance, resistance, medium resistance, susceptible and highly susceptible. This can more accurately reflect the real resistance differences of cruciferous vegetable materials to diamondback moth and has significant guiding significance for insect resistance breeding.

[0038] (10) This invention uses this device to identify insect-resistant germplasm resources of cruciferous vegetables, filling the gap in the identification device and technical method for insect-resistant germplasm resources of the diamondback moth, a major pest of cruciferous vegetables. It has pioneered the creation of an insect-resistant germplasm resource identification device and method for cruciferous vegetables. This method is simple to operate, low in cost, and can accurately detect the susceptibility and high susceptibility of cruciferous vegetables to the diamondback moth. During the experiment, the interference of external factors on the experimental results is avoided, which greatly restores the real situation in the field and improves the authenticity and accuracy of the experimental results. It saves time and effort, has good repeatability, and further ensures the reliability and stability of the experimental data, filling the gap in the current lack of devices and methods for evaluating and identifying the insect resistance of cruciferous vegetables in insect-resistant breeding. Attached Figure Description

[0039] Figure 1 This is a schematic diagram of the device for evaluating and identifying the insect resistance of cruciferous vegetables in this invention;

[0040] Figure 2 This is a physical image of the device for evaluating and identifying the insect resistance of cruciferous vegetables in this invention;

[0041] Figure 3 This is a photograph of the petri dish and the aeration mesh used in this invention.

[0042] Figure 4 This is a first-view image showing the device for evaluating and identifying the insect resistance of cruciferous vegetables in this invention in its operational state.

[0043] Figure 5 This is a second-view diagram showing the usage status of the device for evaluating and identifying the insect resistance of cruciferous vegetables in this invention;

[0044] Figure 6 This is a schematic diagram illustrating the experimental implementation of the device for evaluating and identifying the insect resistance of cruciferous vegetables in this invention;

[0045] Figure 7 In the example, the mortality rate of diamondback moth larvae after feeding on cruciferous vegetables such as cabbage and broccoli is given, where A is the mortality rate of diamondback moth larvae after feeding on cabbage and B is the mortality rate of diamondback moth larvae after feeding on broccoli.

[0046] Figure 8 In the examples, the weight change rate of diamondback moth larvae after feeding on cruciferous vegetables such as cabbage and broccoli is given, where A is the weight change rate of diamondback moth larvae after feeding on cabbage and B is the weight change rate of diamondback moth larvae after feeding on broccoli.

[0047] Figure 9In the examples, the leaf damage rates of diamondback moth larvae feeding on cruciferous vegetables such as cabbage and broccoli are shown. A represents the leaf damage rate of diamondback moth larvae feeding on cabbage varieties, and B represents the leaf damage rate of diamondback moth larvae feeding on broccoli varieties.

[0048] Figure 10 In this example, the resistance levels of diamondback moth larvae feeding on cruciferous vegetables such as cabbage and broccoli are used, where A is the resistance level of diamondback moth larvae feeding on cabbage varieties and B is the resistance level of diamondback moth larvae feeding on broccoli varieties.

[0049] In the picture: 1. Breathable mesh; 2. Lid; 3. Diamondback moth 3rd instar larva; 4. Leaf; 5. Moistened filter paper; 6. Moisturizing cotton; 7. Plastic petri dish; 8. 2% agarose. Detailed Implementation

[0050] The present invention will be further described below with reference to embodiments, but these embodiments are not intended to limit the scope of the invention.

[0051] like Figure 1-3 As shown: The apparatus for identifying insect-resistant germplasm resources of cruciferous vegetable crops according to the present invention includes a plastic petri dish 7, 2% agarose 8, moistened filter paper 5, moisture-retaining cotton 6, a lid 2, and a breathable mesh 1; the bottom of the plastic petri dish 7 contains 2% agarose 8; the moistened filter paper 5 is placed on the 2% agarose 8; the breathable mesh 1 is provided on the lid 2, and the plastic petri dish 7, 2% agarose 8, moistened filter paper 5, breathable mesh 1, and lid 2 form a sealed space, and diamondback moth larvae 3 and leaves 4 are placed in the plastic petri dish 7; the petiole of the leaves 4 is wrapped with moisture-retaining cotton 6.

[0052] In this embodiment, the plastic petri dish 7 has a diameter of 90 mm and a height of 20 mm, the aeration mesh 1 has a mesh size of 100, the filter paper 5 has a diameter of 86 mm, and the agarose concentration is 2%.

[0053] Please see Figure 4-6 The present invention provides a method for identifying insect-resistant germplasm resources of cruciferous vegetable crops, comprising the following:

[0054] Step S1: Select the cruciferous vegetable material to be identified;

[0055] Select the cruciferous vegetable varieties to be identified, and sow different seeds in culture pots to cultivate seedlings. In this example, the culture pots are 11 cm in top diameter, 7 cm in bottom diameter, and 8 cm in height. Sow 3 seeds in each culture pot to ensure that there are seedlings in each pot. Then place them in a greenhouse. When the cruciferous vegetables grow to 4 leaves, remove the weak seedlings and keep the healthy and strong seedlings. Then, when the seedlings grow to 8 leaves, take the 4th-5th leaves for insect resistance identification. Repeat 3 times for each variety.

[0056] Step S2: Preparation of evaluation and identification equipment;

[0057] Select a 90 mm diameter plastic petri dish. Place 2% agarose at the bottom of the petri dish and place an 86 mm diameter filter paper on top of the agarose. Use a brush to moisten the filter paper with sterile water. Cut out the central 40-60 mm area of ​​the petri dish lid and use hot melt glue to attach a 100-mesh aeration mesh to the empty area.

[0058] Step S3: Inoculate the leaves of the cruciferous vegetables with third-instar larvae of the diamondback moth;

[0059] Select vigorous third-instar larvae of the diamondback moth. Use a soft brush to pick up the larvae and gently place them on the leaves to be tested in a petri dish. Cover the petri dish with absorbent paper and then put on the lid. In this example, 10 third-instar larvae of the diamondback moth are inoculated into each petri dish. The petri dishes are then placed in an artificial climate insect rearing room with an ambient temperature of 25 ± 1℃ and a relative humidity of 65%.

[0060] Step S4: Observation of the survival and development of diamondback moth larvae;

[0061] At 24 h, 48 h, and 72 h after inoculation, the culture apparatus was turned on sequentially, and the diamondback moth larvae in the petri dishes were observed and recorded.

[0062] ① The number of diamondback moth larvae that died at each time point;

[0063] ② The weight changes of surviving larvae, weigh them and calculate the average weight;

[0064] ③ Leaf damage: The total damaged area and total leaf area are obtained by measuring the area of ​​leaf damage caused by larvae feeding.

[0065] The above-mentioned data were used to characterize the survival ability, growth and development status, and feeding damage of diamondback moth larvae on different cruciferous vegetable materials.

[0066] Step S5: Data analysis of resistance of cruciferous vegetable crops to diamondback moth;

[0067] After obtaining the above raw data, the following resistance evaluation indicators were calculated respectively:

[0068] 1) The corrected mortality rate M of larvae, expressed as a percentage, is calculated using the following formula:

[0069] in, To address the mortality rate of diamondback moth larvae in the treatment group, The mortality rate of diamondback moth larvae in the control group is shown in percentages (%).

[0070] 2) The larval weight change rate P, expressed as a percentage, is calculated using the following formula:

[0071] in, The average weight of surviving larvae in the treatment group. The average weight of surviving larvae in the control group is given in milligrams (mg).

[0072] 3) The leaf damage rate L, expressed as a percentage, is calculated using the following formula:

[0073] in, This represents the total area of ​​leaf damage caused by the larvae feeding. The total area of ​​the leaves is expressed in square centimeters (cm). 2 ).

[0074] After obtaining the above three indicators, a weighted calculation is performed according to their relative weights to obtain the comprehensive insect resistance index I of cruciferous vegetable materials. The calculation formula is as follows:

[0075] Wherein, I is the comprehensive insect resistance index, ranging from 0 to 100; M is the corrected mortality rate, in percentage (%); P is the surviving larval weight change rate, in percentage (%); and L is the leaf damage rate, in percentage (%). The comprehensive insect resistance index I is positively correlated with larval mortality and weight change rate, and negatively correlated with leaf damage rate.

[0076] Based on the numerical range of the comprehensive insect resistance index I, cruciferous vegetable materials are divided into different resistance levels, and the specific grading standards are shown in Table 1:

[0077]

[0078] This allows for a comprehensive evaluation of the diamondback moth resistance levels of different cruciferous vegetable materials, enabling the screening of highly resistant and susceptible materials.

[0079] This embodiment selected 10 cabbage varieties and 10 broccoli varieties as test materials. The cabbage variety JingFeng No.1, a major field-grown variety in my country, was used as the control, and the broccoli variety ZhongQing No.15, also a major field-grown variety in my country, was used as the control. The materials were divided into group A (cabbage) and group B (broccoli) for inoculation, observation, and identification. A resistance grading chart was drawn based on the calculated comprehensive insect resistance index I, as shown below. Figures 7 to 10As shown in Table 1, A represents the resistance level of 10 cabbage varieties to diamondback moth larvae, and B represents the resistance level of 10 broccoli varieties to diamondback moth larvae. The corresponding resistance levels and comprehensive resistance indices are listed in Table 2. Table 2 shows the comprehensive resistance index and resistance level statistics of diamondback moth larvae after feeding on cabbage. Table 3 shows the mortality rate, surviving larval weight change rate, and leaf damage rate of diamondback moth larvae after feeding on broccoli. Table 4 shows the comprehensive resistance index and resistance level statistics of diamondback moth larvae after feeding on broccoli.

[0080] The experimental results show that the device and method established in this invention can clearly distinguish the differences in resistance to diamondback moth among different cruciferous vegetable materials, and have good stability and repeatability.

[0081] Table 1 Mortality rate, weight change rate, and leaf damage rate of diamondback moth larvae after feeding on cabbage.

[0082]

[0083] Table 2. Statistics on the comprehensive resistance index and resistance level of diamondback moth larvae after feeding on cabbage.

[0084]

[0085] Table 3 Mortality rate, surviving larval weight change rate, and leaf damage rate of diamondback moth larvae after feeding on broccoli.

[0086]

[0087] Table 4. Statistics on the comprehensive resistance index and resistance level of diamondback moth larvae after feeding on broccoli.

[0088]

[0089] The foregoing has shown and described the basic principles, main features, and advantages of the present invention. However, the above description is merely a specific embodiment of the present invention, and the technical features of the present invention are not limited thereto. Any other embodiments derived by those skilled in the art without departing from the technical solution of the present invention should be covered within the patent scope of the present invention.

Claims

1. An identification device for diamondback moth-resistant germplasm resources of cruciferous vegetable crops, characterized in that, The system includes a breathable mesh (1), a lid (2), moistened filter paper (5), moistening cotton (6), a petri dish (7), and a 2% agarose substrate (8). The lid (2) is provided with a breathable mesh (1). The bottom of the petri dish (7) is covered with a 2% agarose substrate (8), and a moistened filter paper (5) is placed on the 2% agarose substrate (8). The petri dish (7), the 2% agarose (8), the moistened filter paper (5), the breathable mesh (1), and the lid (2) form a closed and breathable culture space. The petri dish (7) is used to place diamondback moth 3rd instar larvae (3) and leaves (4) for feeding. The petioles of the leaves (4) are wrapped with moistening cotton (6).

2. The identification device for diamondback moth-resistant germplasm resources of cruciferous vegetable crops according to claim 1, characterized in that, The mesh number of the breathable mesh (1) is not less than 100 mesh.

3. The identification device for diamondback moth-resistant germplasm resources of cruciferous vegetable crops according to claim 1, characterized in that, The moisturizing cotton is moistened with sterile water.

4. A method for identifying diamondback moth-resistant germplasm resources of cruciferous vegetable crops according to any one of claims 1-3, characterized in that, The method is as follows: Step S1: Select cruciferous vegetable materials to be identified; Step S2: Inoculate the third instar larvae of the diamondback moth onto the leaves of the cruciferous vegetables; Step S3: Cultivate the inoculated diamondback moth larvae; Step S4: Observe and record the survival and development status of diamondback moth larvae; Step S5: Conduct a comprehensive evaluation and data analysis of the diamondback moth resistance of cruciferous vegetable crop materials.

5. The method for identifying diamondback moth-resistant germplasm resources of cruciferous vegetable crops according to claim 4, characterized in that, The specific steps for selecting cruciferous vegetable materials to be identified in step S1 are as follows: Sow different cruciferous vegetable seeds in culture pots, with 3 seeds sown in each culture pot, and then place them in a greenhouse. When the cruciferous vegetables grow to 4 leaves, remove the weak seedlings and keep the healthy and strong seedlings. Then, when the seedlings grow to 8 leaves, take the 4th-5th leaves for insect resistance identification. Repeat this process 3 times for each variety.

6. The method for identifying diamondback moth-resistant germplasm resources of cruciferous vegetable crops according to claim 4, characterized in that, The preparation of the evaluation and identification device described in step S2 is as follows: Select a plastic petri dish with a diameter of 90 mm, lay a 2% agarose matrix at the bottom of the petri dish, place a filter paper with a diameter of 86 mm on top of the agarose, moisten the filter paper with sterile water using a brush, cut out the central area of ​​40-60 mm of the petri dish lid, and use hot melt glue to attach a 100-mesh breathable mesh to the empty area.

7. The method for identifying diamondback moth-resistant germplasm resources of cruciferous vegetable crops according to claim 4, characterized in that, The inoculation of third-instar larvae of the diamondback moth in step S2 is carried out as follows: Select healthy and active third-instar larvae of the diamondback moth, and inoculate each petri dish with a predetermined number of third-instar larvae, preferably 10. Cover the top of the petri dish with absorbent paper, and then put the petri dish lid on.

8. The method for identifying diamondback moth-resistant germplasm resources of cruciferous vegetable crops according to claim 4, characterized in that, Step S3: Culture conditions after inoculation of diamondback moth 3rd instar larvae: Cultured in an artificial climate insect rearing room with an ambient temperature of 25 ± 1℃ and a relative humidity of 65%.

9. The method for identifying diamondback moth-resistant germplasm resources of cruciferous vegetable crops according to claim 4, characterized in that, The observation of the survival and development of diamondback moth described in step S4 is as follows: 24 h, 48 h and 72 h after completing step S3, the diamondback moth larvae in the petri dish are observed, and the number of dead diamondback moth larvae, the weight change of surviving larvae and the damaged area of ​​the leaves on cruciferous vegetable leaves are recorded.

10. The method for identifying diamondback moth-resistant germplasm resources of cruciferous vegetable crops according to claim 4, characterized in that, In step S5, a comprehensive evaluation of the diamondback moth resistance of cruciferous vegetable materials is conducted, specifically including: Step S5.1: Calculate the corrected mortality rate, using the following formula: ; in, To address the mortality rate of diamondback moth larvae in the treatment group, The mortality rate of diamondback moth larvae in the control group is shown in percentage (%). The corrected mortality rate is used to eliminate the influence of external environment or experimental error. Step S5.2: Calculate the rate of change in surviving larvae body weight by measuring the average body weight of surviving larvae in the experimental and control groups. The formula is: ; in, The average weight of surviving larvae in the treatment group. The average weight of surviving larvae in the control group is shown in milligrams (mg). Changes in body weight can reflect the inhibitory effect of plants on the growth and development of diamondback moth. Step S5.3: Calculate the blade damage rate using the following formula: ; in, This represents the total area of ​​leaf damage caused by the larvae feeding. The total area of ​​the leaves is expressed in square centimeters (cm). 2 The degree of damage caused by larval feeding is quantified by calculating the ratio of damaged leaf area to total leaf area. Step S5.4: Assign weights to the above three indicators and calculate the comprehensive insect resistance index I through weighted average. The formula is: ; Wherein, I is the comprehensive insect resistance index, ranging from 0 to 100; M is the corrected mortality rate, in percentage (%); P is the surviving larval weight change rate, in percentage (%); and L is the leaf damage rate, in percentage (%). The comprehensive insect resistance index I is positively correlated with larval mortality and weight change rate, and negatively correlated with leaf damage rate. A weighted method is used to comprehensively consider mortality, weight change, and leaf damage to avoid the bias of a single indicator. Step S5.5: Based on the numerical range of the comprehensive insect resistance index I, cruciferous vegetable materials are divided into different resistance levels. The specific grading standards are shown in Table 1: ； Based on the number of diamondback moth larvae that died, the weight changes of surviving larvae, and the area of ​​leaf damage recorded in step S4, the corrected mortality rate of larvae, the weight change rate of surviving larvae, and the leaf damage rate were determined respectively. Further, the above indicators were weighted according to a preset weight ratio to obtain the comprehensive insect resistance index I of the cruciferous vegetable materials. The comprehensive insect resistance index is positively correlated with larval mortality and weight changes, and negatively correlated with the degree of leaf damage. Based on the numerical range of the comprehensive insect resistance index I, the cruciferous vegetable materials were divided into five resistance levels: highly resistant, resistant, moderately resistant, susceptible, and highly susceptible, thereby achieving the identification of diamondback moth resistant germplasm resources in cruciferous vegetables.

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