SSR primer combination for cedrela sinensis genetic diversity detection and application thereof

Abstract

The invention discloses an SSR primer combination for cedrela sinensis genetic diversity detection and application of the SSR primer combination. The SSR primer combination contains 13 pairs of SSR primer pairs which are used for carrying out specific amplification on SSR sites TS01-TS013 in the DNA (Deoxyribose Nucleic Acid) of the Toona sinense genome of the toona sinensis; the SSR primer combination is applied to cedrela sinensis genetic diversity detection, cedrela sinensis core germplasm population construction or cedrela sinensis variety identification. The SSR primer combination provided by the invention can accurately and efficiently detect the genetic diversity of the cedrela sinensis, provides a powerful tool for constructing a cedrela sinensis core germplasm population, and lays a solid foundation for long-term breeding and resource management of the cedrela sinensis.

Description

Technical Field

[0001] This invention relates to the field of genetic diversity detection technology. Specifically, it relates to SSR primer combinations for detecting genetic diversity in Toona sinensis and their applications. Background Technology

[0002] The construction of core germplasm aims to remove redundant individuals from a population, representing the genetic diversity of the entire population to the greatest extent possible with minimal genetic resources, thereby significantly improving the management and utilization efficiency of plant germplasm resources. Currently, domestic and international research mainly focuses on the construction of core germplasm for crops, but reports on the construction of core germplasm for forest trees are relatively rare.

[0003] Chinese toon (Toona sinensis) is a tree species belonging to the genus Toona in the family Meliaceae. Due to its excellent timber quality and rapid growth, it is known as "Chinese mahogany." Modern scientific research has proven that Chinese toon sprouts are rich in nutrients and medicinal components such as protein, vitamins, trace elements, polyphenols, flavonoids, and quercetin. It is a high-value forest-grown health vegetable with various effects including antioxidant, antibacterial, blood sugar-lowering, lipid-lowering, and anti-cancer properties. Because Chinese toon has value as timber, vegetable, and medicinal material, it plays an important role in promoting national health and increasing farmers' income.

[0004] Developing tools for detecting genetic diversity in Toona sinensis is of great significance for better research and utilization of its genetic resources. SSR (microsatellite sequence) markers are an effective tool for detecting genetic diversity. Therefore, it is urgent to develop primer combinations for amplifying SSR sequences in Toona sinensis and to use these primer combinations for detecting genetic diversity and constructing core germplasm populations, so as to achieve comprehensive detection and scientific utilization of Toona sinensis's genetic diversity. Summary of the Invention

[0005] Therefore, the technical problem to be solved by this invention is to provide an SSR primer combination for detecting the genetic diversity of Toona sinensis and its application. These primer combinations can accurately and efficiently detect the genetic diversity of Toona sinensis, providing a powerful tool for constructing a core germplasm population and laying a solid foundation for the long-term breeding and resource management of Toona sinensis.

[0006] To solve the above-mentioned technical problems, the present invention provides the following technical solution:

[0007] This SSR primer set for detecting genetic diversity in Toona sinensis contains 13 pairs of SSR primers for specifically amplifying SSR loci TS01–TS013 in the Toona sinensis genomic DNA. Specifically: the SSR primer pair for specific amplification of the TS01 locus consists of primers with sequences shown in SEQ ID NO. 1 and SEQ ID NO. 2, respectively; the SSR primer pair for specific amplification of the TS02 locus consists of primers with sequences shown in SEQ ID NO. 3 and SEQ ID NO. 4, respectively; the SSR primer pair for specific amplification of the TS03 locus consists of primers with sequences shown in SEQ ID NO. 5 and SEQ ID NO. 6, respectively; the SSR primer pair for specific amplification of the TS04 locus consists of primers with sequences shown in SEQ ID NO. 7 and SEQ ID NO. 8, respectively; and the SSR primer pair for specific amplification of the TS05 locus consists of primers with sequences shown in SEQ ID NO. 9 and SEQ ID NO. 1, respectively. The primer pair shown in NO. 10 consists of primers for specific amplification of the TS06 site, and consists of primers with sequences shown in SEQ ID NO. 11 and SEQ ID NO. 12, respectively; the SSR primer pair for specific amplification of the TS07 site consists of primers with sequences shown in SEQ ID NO. 13 and SEQ ID NO. 14, respectively; the SSR primer pair for specific amplification of the TS08 site consists of primers with sequences shown in SEQ ID NO. 15 and SEQ ID NO. 16, respectively; the SSR primer pair for specific amplification of the TS09 site consists of primers with sequences shown in SEQ ID NO. 17 and SEQ ID NO. 18, respectively; the SSR primer pair for specific amplification of the TS010 site consists of primers with sequences shown in SEQ ID NO. 19 and SEQ ID NO. 20, respectively; and the SSR primer pair for specific amplification of the TS011 site consists of primers with sequences shown in SEQ ID NO. 21 and SEQ ID NO. 12, respectively. The primer pair shown in NO.22 consists of primers with sequences as shown in SEQ ID NO.23 and SEQ ID NO.24, respectively; the SSR primer pair shown in SEQ ID NO.25 and SEQ ID NO.26 consists of primers with sequences as shown in SEQ ID NO.25 and SEQ ID NO.26, respectively.

[0008] Applications of SSR primer combinations for detecting genetic diversity in Toona sinensis: The application includes using the aforementioned SSR primer combinations for detecting genetic diversity in Toona sinensis; or, using the aforementioned SSR primer combinations to construct a core germplasm population of Toona sinensis; or, using the aforementioned SSR primer combinations for the identification of Toona sinensis varieties.

[0009] The above-mentioned application, the method for detecting the genetic diversity of Toona sinensis includes the following steps: (1) extracting genomic DNA from Toona sinensis; (2) performing PCR amplification on the genomic DNA extracted in step (1) using the above-mentioned SSR primer combination; (3) performing electrophoresis detection on the PCR product obtained in step (2), and analyzing the genetic diversity of Toona sinensis based on the electrophoresis results. Using SSR primers, the SSR sites in the genomic DNA of Toona sinensis are amplified by PCR amplification, and the electrophoresis results of the PCR products are analyzed. The genetic diversity of the species can be quantitatively assessed by information such as the number and size differences of bands and heterozygosity.

[0010] The above application involves constructing a core germplasm population of Toona sinensis, following these steps: S1. Select candidate plants from the wild Toona sinensis population and collect seeds from individual plants; S2. Cultivate individuals from the seeds of the candidate plants collected in step S1, with individuals developed from the seeds of the same candidate plant forming a candidate family; S3. Select 1-5 individuals from each candidate family, and randomly select one individual from each existing Toona sinensis variety, adding these individuals to form a basic population; extract genomic DNA from each individual in the basic population; S4. Using the above-mentioned SSR primer combination, perform SSR on the genes extracted in step S3 on individual plants. The DNA was amplified by PCR, and the PCR products were detected by electrophoresis. S5: Based on the electrophoresis results, the genetic diversity index of the basic population and the genetic distance between individuals in the basic population were analyzed. Individuals were selected from the basic population according to different sampling ratios, and the individuals selected at each sampling ratio formed a candidate core germplasm population. S6: The capture rate of the genetic diversity index of the basic population for each candidate core germplasm population was calculated, and the sampling ratio / capture rate ratio of each candidate core germplasm population was calculated. The candidate core germplasm population with a capture rate greater than or equal to 90% and the smallest sampling ratio / capture rate ratio was selected as the core germplasm population. Randomly selecting one individual from each existing Toona sinensis variety, along with a specific single plant (a single plant with a special trait), and adding these individuals to the basic population is beneficial for improving the genetic diversity of the basic population.

[0011] In the above application, in step S1, when selecting candidate plants from each wild Toona sinensis population, the distance between any two candidate plants must be greater than or equal to 250 m; the number of candidate plants selected from each wild Toona sinensis population must be greater than or equal to 10; the selection criteria for candidate plants are: straight trunk, branch height greater than 8.0 m, heartwood percentage greater than 80%, and heartwood color reddish-brown. When selecting candidate plants from a wild Toona sinensis population, if the distance between two individuals is less than 250 m, the two Toona sinensis plants are generally closely related and have little genetic difference, meaning that these two individuals should not be selected as candidate plants simultaneously.

[0012] In the above application, in step S3, one individual is randomly selected from each candidate family to form a basic population; or, two or more individuals are selected from each candidate family to form a basic population, and the maximum difference in plant height between the two or more selected individuals is greater than or equal to 10cm, the maximum difference in ground diameter is greater than or equal to 0.2cm, and there are differences in the color of the buds and the shape of the leaves among the selected individuals.

[0013] In the above application, in step S5, the genetic diversity index includes the number of alleles, the effective number of alleles, observed heterozygosity, expected heterozygosity, Shannon information index, and polymorphism information index; among which, the expected heterozygosity of the basic population is greater than 0.8, and the Shannon information index is greater than 1.8. When the expected heterozygosity of the basic population is less than or equal to 0.8, and the Shannon information index is less than or equal to 1.8, the genetic diversity of the basic population is low and cannot meet the requirements for constructing a core germplasm population; therefore, the basic population should be reconstructed.

[0014] In the above application, step S5 involves selecting individuals from the base population at sampling ratios of 5%, 10%, 15%, 20%, 25%, and 30%, respectively, to form six candidate core germplasm populations. The sampling ratio of the candidate core germplasm population refers to the percentage of individuals in the candidate core germplasm population relative to the total number of individuals in the base population. A larger sampling ratio results in an excessive number of individuals in the candidate core germplasm populations, which is detrimental to management. A sampling ratio gradient of 5% provides a reasonable difference between the number of individuals in each candidate core germplasm population and the capture rate of the genetic diversity index.

[0015] In step S5, the sampling strategy for selecting individuals from the base population is to maximize the number of alleles (genetic diversity level), and this step is automatically completed by the Core Hunter 3 software. That is, given a fixed sampling ratio, when selecting individuals from the base population to construct a candidate core germplasm population, the goal is to intentionally include as many alleles as possible in the candidate core germplasm population to ensure richer genetic diversity. This is beneficial for achieving a higher capture rate of the genetic diversity index in the final selected core germplasm population and minimizing the genetic diversity difference between it and the base population.

[0016] In step S6, the capture rate of the genetic diversity index of the basic population by each candidate core germplasm population should be compared. Among the candidate core germplasm populations with higher capture rates of genetic diversity index, the smaller candidate core germplasm population (i.e., the candidate core germplasm population with a smaller sampling ratio) should be selected as the final core germplasm population. Meanwhile, the capture rate of the genetic diversity index of the basic population by the final selected core germplasm population should not be less than 90%, so that the core germplasm population is more representative.

[0017] As a preferred technical solution, the capture rate of Shannon information index, expected heterozygosity, or number of alleles is selected as the capture rate of the genetic diversity index of the base population for each candidate core germplasm population. Shannon information index can comprehensively consider the number of species and the evenness of individual distribution, and is suitable as a representative of the genetic diversity index capture rate.

[0018] In the above application, in step S4, when performing PCR amplification on the genomic DNA of Toona sinensis, the reaction system volume used is 20 μL, including 10 μL of 2× PCR buffer, 0.4 μL of dNTP, 0.5 μL of F primer, 0.5 μL of R primer, 1 μL of template DNA, and 0.2 μL of Taq DNA polymerase, with the remainder being ultrapure water.

[0019] In the above application, when performing PCR amplification of the genomic DNA of Toona sinensis in step S4, the reaction program used is as follows: 94℃ pre-denaturation for 4 min; 94℃ denaturation for 1 min, 52-55℃ annealing for 0.5 min, 72℃ extension for 1 min, 35 cycles; 72℃ final extension for 10 min.

[0020] The technical solution of the present invention achieves the following beneficial technical effects:

[0021] 1. This invention provides a specific SSR primer combination for Toona sinensis, which, through PCR amplification and electrophoresis detection, can rapidly and accurately identify genetic differences among individual Toona sinensis plants. Using this SSR primer combination, a genetically diverse and highly representative core germplasm population of Toona sinensis can be effectively constructed, while reducing the number of redundant individuals. This significantly improves the management and utilization efficiency of Toona sinensis germplasm resources, enhances the efficiency and accuracy of Toona sinensis genetic diversity research, and lays a solid foundation for subsequent genetic analysis and breeding work.

[0022] 2. The SSR primer combinations of this invention not only play an important role in the construction of core germplasm populations, but can also be widely applied in fields such as genetic diversity assessment and variety identification of Toona sinensis populations. By improving the management efficiency and utilization effect of germplasm resources, this invention helps protect the genetic diversity of Toona sinensis, prevent resource loss and degradation, promote the sustainable utilization of Toona sinensis resources, and contribute to national health and increased farmers' income. Attached Figure Description

[0023] Figure 1 In this embodiment of the invention, the genetic diversity index capture rate of each candidate core germplasm population was compared when the SSR primer composition was used to construct the core germplasm population of Toona sinensis.

[0024] Figure 2 The technical flowchart of using SSR primer composition for constructing core germplasm population of Toona sinensis in this invention. Detailed Implementation

[0025] like Figure 2 This is a flowchart illustrating the technical process of this embodiment. The inventors conducted a survey of wild Toona sinensis germplasm resources, completing the collection of germplasm from all distribution areas of Toona sinensis, and collecting germplasm resources from 32 wild Toona sinensis populations. In addition, existing Toona sinensis varieties and unique individual plants from both domestic and international sources were collected. All of the above germplasm resources were planted in Kaihua County, Zhejiang Province.

[0026] For the collected wild Toona sinensis populations, at least 10 candidate trees should be selected from each population for seed collection, with a spacing of 250m or more between any two candidate trees. When selecting candidate trees, the criteria are: straight trunk, branch height greater than 8.0m, heartwood content greater than 80%, and heartwood color reddish-brown.

[0027] Seeds from collected candidate plants are cultured into individuals. A single candidate plant typically produces multiple seeds, and individuals developed from seeds from the same candidate plant constitute a candidate family.

[0028] Based on the differences in individual growth status and bud color, and adhering to the principle of maximizing differences in growth status while also exhibiting significant differences in bud color and leaf shape, 1 to 5 individuals were selected from each candidate family, for a total of 1020 individuals. When selecting one individual from each candidate family, the selection was random. When selecting two or more individuals from each candidate family, the maximum difference in plant height between the selected individuals must be greater than or equal to 10 cm, the maximum difference in ground diameter must be greater than or equal to 0.2 cm, and there must be differences in bud color and leaf shape.

[0029] In addition, one individual was randomly selected from each of the five existing Toona sinensis varieties to be added to the basic population, and 15 individual plants with special traits were also added to the basic population (a total of 20 individuals were selected from the existing Toona sinensis varieties and the individual plants with special traits). These 1040 individuals constituted the basic population. All individuals in the basic population were numbered, and genomic DNA was extracted from each individual.

[0030] Based on whole-genome and transcriptome sequencing data from *Toona sinensis*, the inventors designed 100 pairs of SSR primers. Through extensive testing and optimization, 13 pairs of SSR primers were selected, capable of specifically amplifying 13 SSR sites in the *Toona sinensis* genomic DNA. Actual testing showed that these 13 pairs of SSR primers yielded products with high polymorphism when used for PCR of *Toona sinensis* genomic DNA. The sequences of these 13 pairs of SSR primers are shown in Table 1.

[0031] Table 1

[0032]

[0033] When using these 13 SSR primer pairs to perform PCR amplification of Toona sinensis genomic DNA from individual plants, one SSR primer pair was added to each reaction system, meaning that each primer in each reaction system specifically amplifies only one SSR site. The PCR reaction system volume was 20 μL, including 10 μL of 2× PCR buffer, 0.4 μL of dNTPs, 0.5 μL of F primer, 0.5 μL of R primer, 1 μL of template DNA, and 0.2 μL of Taq DNA polymerase, with the remainder being ultrapure water.

[0034] The reaction program was as follows: pre-denaturation at 94℃ for 4 min, 35 denaturation-annealing cycles (denaturation at 94℃ for 1 min, annealing at 52-55℃ for 0.5 min, extension at 72℃ for 1 min), and final extension at 72℃ for 10 min.

[0035] After PCR, the PCR products were subjected to electrophoresis, and the results were analyzed using GenAlex v6.5 software to determine the genetic diversity index of the basic population. The indicators for detecting genetic diversity included the number of alleles, the effective number of alleles, the Shannon information index, observed heterozygosity, expected heterozygosity, and polymorphism information index. When the expected heterozygosity of the basic population was less than or equal to 0.8, and the Shannon information index was less than or equal to 1.8, the genetic diversity of the basic population was considered low and did not meet the requirements for constructing a core germplasm population; therefore, the basic population should be reconstructed.

[0036] Six candidate core germplasm populations were obtained by selecting individuals using Core Hunter 3 and Powermarker software at sampling ratios of 5%, 10%, 15%, 20%, 25%, and 30%, respectively. The selection strategy was to maximize the number of alleles (genetic diversity). That is, given a fixed sampling ratio, when selecting individuals from the base population to form candidate core germplasm populations, the aim was to intentionally include as many alleles as possible in these populations. This ensures a higher capture rate of the genetic diversity index in the final selected core germplasm populations and a smaller difference in genetic diversity between them and the base population.

[0037] The differences in the capture rates of genetic diversity indices (including the number of alleles, the number of effective alleles, observed heterozygosity, expected heterozygosity, Shannon information index, and polymorphism information index) between each candidate core germplasm population and the base population were compared to assess the representativeness of each candidate core germplasm population. Specifically, the final core germplasm population should capture as high a rate of genetic diversity indices of the base population as possible. Furthermore, the number of individuals in the core germplasm population should be as small as possible, meaning the core germplasm population should contain as many variant types as possible from the base population, while maintaining a small size to facilitate management and breeding operations.

[0038] Table 2 shows the genetic diversity index of the basic population (the average value of each locus is used as the final calculation result).

[0039] Table 2 Genetic diversity index of the basic population of Toona sinensis

[0040]

[0041] The abbreviations in the table represent the following: Na: number of alleles; Ne: number of effective alleles; I: Shannon's information index; Ho: observed heterozygosity; He: expected heterozygosity; PIC: polymorphic information content.

[0042] like Figure 1 This section compares the capture rates of genetic diversity indices for candidate core germplasm populations using different sampling strategies. The horizontal axis represents the sampling percentages of 5%, 10%, 15%, 20%, 25%, and 30%, respectively. CH and PM represent the Core Hunter3 and Powermarker methods, respectively, and the numbers following CH and PM represent the number of individuals in the corresponding candidate core germplasm populations.

[0043] Depend on Figure 1 It can be seen that when Core Hunter 3 software is used for sampling, the sampling ratio is 20%, the number of alleles, the number of effective alleles and the expected heterozygosity capture rate of the candidate core germplasm population are relatively high, and more than 95% of the genetic diversity of the basic population can be captured.

[0044] The capture rate of the genetic diversity index of each candidate core germplasm population relative to the base population is calculated. Since the genetic diversity index includes multiple indices, in this embodiment, the capture rate of the Shannon information index is used as the capture rate of the genetic diversity index of the candidate core germplasm population relative to the base population. In other embodiments, the capture rate of one of the genetic diversity indices can be selected as the capture rate of the genetic diversity index of the candidate core germplasm population relative to the base population, depending on actual needs. Then, the sampling ratio / capture rate of each candidate core germplasm population is calculated, and the candidate core germplasm population with a capture rate greater than or equal to 90% and the smallest sampling ratio / capture rate is selected as the core germplasm population.

[0045] Finally, the candidate core germplasm population selected using Core Hunter 3 software, representing 20% ​​of the total sample, was chosen as the final core germplasm population. This core germplasm population contains 208 individual Toona sinensis plants. Struttre analysis of this core germplasm population revealed that it covers the entire distribution area of ​​Toona sinensis (i.e., in all wild Toona sinensis populations collected, at least one candidate individual's offspring from each population was selected into this core germplasm population). Phenotypic traits such as seedling height, compound leaf length, number of compound leaves, number of branches, and number of hairs on the underside of leaves in this core germplasm population were measured (as shown in Table 3), demonstrating that the core germplasm population possesses rich phenotypic variation.

[0046] Table 3. Statistical table of phenotypic traits of core germplasm

[0047]

[0048] Besides being used for constructing core germplasm populations, the SSR primer combinations shown in Table 1 can be used to detect the genetic diversity of individual or population *Toona sinensis* plants to assess the level of genetic diversity in a particular individual or population. Additionally, the SSR primer combinations shown in Table 1 can also be used for the identification of *Toona sinensis* varieties. By comparing the SSR genotyping of the plant to be identified with that of known varieties at the aforementioned 13 loci, the variety of the plant to be identified can be determined.

[0049] Obviously, the above embodiments are merely illustrative examples for clear explanation and are not intended to limit the implementation. Those skilled in the art will recognize that other variations or modifications can be made based on the above description. It is neither necessary nor possible to exhaustively list all possible implementations here. However, obvious variations or modifications derived therefrom are still within the scope of protection of the claims of this patent application.

Claims

1. An SSR primer combination for detecting genetic diversity in Toona sinensis, characterized in that, This SSR primer set contains 13 pairs of SSR primers for specific amplification of SSR sites TS01–TS013 in the genomic DNA of *Toona sinensis*, including: The SSR primer pair used for specific amplification of the TS01 site consists of primers with sequences as shown in SEQ ID NO.1 and SEQ ID NO.2, respectively; The SSR primer pair used for specific amplification of the TS02 site consists of primers with sequences as shown in SEQ ID NO.3 and SEQ ID NO.4, respectively; The SSR primer pair used for specific amplification of the TS03 site consists of primers with sequences as shown in SEQ ID NO.5 and SEQ ID NO.6, respectively; The SSR primer pair used for specific amplification of the TS04 site consists of primers with sequences shown in SEQ ID NO.7 and SEQ ID NO.8, respectively; The SSR primer pair used for specific amplification of the TS05 site consists of primers with sequences as shown in SEQ ID NO.9 and SEQ ID NO.10, respectively; The SSR primer pair used for specific amplification of the TS06 site consists of primers with sequences as shown in SEQ ID NO.11 and SEQ ID NO.12, respectively; The SSR primer pair used for specific amplification of the TS07 site consists of primers with sequences as shown in SEQ ID NO.13 and SEQ ID NO.14, respectively; The SSR primer pair used for specific amplification of the TS08 site consists of primers with sequences as shown in SEQ ID NO.15 and SEQ ID NO.16, respectively; The SSR primer pair used for specific amplification of the TS09 site consists of primers with sequences as shown in SEQ ID NO.17 and SEQ ID NO.18, respectively; The SSR primer pair used for specific amplification of the TS010 site consists of primers with sequences as shown in SEQ ID NO.19 and SEQ ID NO.20, respectively; The SSR primer pair used for specific amplification of the TS011 site consists of primers with sequences as shown in SEQ ID NO.21 and SEQ ID NO.22, respectively; The SSR primer pair used for specific amplification of the TS012 site consists of primers with sequences as shown in SEQ ID NO.23 and SEQ ID NO.24, respectively; The SSR primer pair used for specific amplification of the TS013 site consists of primers with sequences shown in SEQ ID NO.25 and SEQ ID NO.26, respectively.

2. The application of SSR primer combinations for detecting genetic diversity in Toona sinensis, characterized in that, The application is to use the SSR primer combination as described in claim 1 for the detection of genetic diversity of Toona sinensis; or, the application is to use the SSR primer combination as described in claim 1 for the construction of a core germplasm population of Toona sinensis; or, the application is to use the SSR primer combination as described in claim 1 for the identification of Toona sinensis varieties.

3. The application according to claim 2, characterized in that, The method for detecting genetic diversity in Toona sinensis includes the following steps: (1) Extracting genomic DNA from Toona sinensis; (2) Perform PCR amplification on the genomic DNA extracted in step (1) using the SSR primer combination as described in claim 1; (3) Perform electrophoresis on the PCR products obtained in step (2) and analyze the genetic diversity of Toona sinensis based on the electrophoresis results.

4. The application according to claim 2, characterized in that, When constructing the core germplasm population of Toona sinensis, follow these steps: S1. Select candidate plants from the wild Chinese toon population and collect seeds from individual plants; S2. The seeds of the candidate plants collected in step S1 are cultivated into individuals. Individuals developed from the seeds of the same candidate plant constitute a candidate family. S3. Select 1 to 5 individuals from each candidate family, and randomly select one individual from each existing Toona sinensis variety, and add the specific single plant to form a basic population; extract the genomic DNA of each individual in the basic population. S4. Using the SSR primer combination as described in claim 1, perform PCR amplification on the genomic DNA extracted in step S3 using individual plants, and detect the PCR products by electrophoresis. S5. Analyze the genetic diversity index of the basic population and the genetic distance between individuals in the basic population based on the electrophoresis results, and select individuals from the basic population according to different sampling ratios. Individuals selected under each sampling ratio form a candidate core germplasm population. S6. Calculate the capture rate of the genetic diversity index of each candidate core germplasm population to the basic population, calculate the sampling ratio / capture rate ratio of each candidate core germplasm population, and select the candidate core germplasm population with a capture rate greater than or equal to 90% and the smallest sampling ratio / capture rate ratio as the selected core germplasm population.

5. The application according to claim 4, characterized in that, In step S1, when selecting candidate trees in each wild Chinese toon population, the distance between any two candidate trees is greater than or equal to 250m; the number of candidate trees selected in each wild Chinese toon population is greater than or equal to 10; the selection criteria for candidate trees are that the trunk is straight, the height of the branches is greater than 8.0m, the heartwood ratio is greater than 80%, and the heartwood color is reddish-brown.

6. The application according to claim 4, characterized in that, In step S3, one individual is randomly selected from each candidate family to form a basic population; or, two or more individuals are selected from each candidate family to form a basic population, and the maximum difference in plant height between the two or more selected individuals is greater than or equal to 10cm, the maximum difference in ground diameter is greater than or equal to 0.2cm, and there are differences in the color of the buds and the shape of the leaves among the selected individuals.

7. The application according to claim 4, characterized in that, In step S5, the genetic diversity index includes the number of alleles, the effective number of alleles, observed heterozygosity, expected heterozygosity, Shannon information index, and polymorphism information index; among which, the expected heterozygosity of the base population is greater than 0.8, and the Shannon information index is greater than 1.

8.

8. The application according to claim 4, characterized in that, In step S5, individuals are selected from the basic population at sampling ratios of 5%, 10%, 15%, 20%, 25%, and 30% respectively, forming six candidate core germplasm populations.

9. The application according to claim 3 or 4, characterized in that, In step S4, when performing PCR amplification on the genomic DNA of Toona sinensis, the reaction system volume used is 20 μL, including 10 μL of 2× PCR buffer, 0.4 μL of dNTP, 0.5 μL of F primer, 0.5 μL of R primer, 1 μL of template DNA, and 0.2 μL of Taq DNA polymerase, with the remainder being ultrapure water.

10. The application according to claim 9, characterized in that, In step S4, the reaction program used for PCR amplification of the genomic DNA of Toona sinensis was as follows: 94℃ pre-denaturation for 4 min; 94℃ denaturation for 1 min, 52-55℃ annealing for 0.5 min, 72℃ extension for 1 min, 35 cycles; 72℃ final extension for 10 min.

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