Screening method and application of macadamia inbred and high-quality germplasm

Abstract

The application provides a screening method and application of macadamia self-compatible and high-quality germplasm, and belongs to the technical field of plant breeding. The application performs bagging self-pollination and natural pollination on the to-be-screened germplasm; performs primary screening based on a self-pollination fruit setting rate of greater than or equal to 0.5%; then determines and compares the self-pollination and natural pollination mature fruit traits of the primary screening compatible germplasm; and through calculation of the self-pollination recession rates of various traits, the germplasm with a self-pollination recession rate of fruit traits lower than a preset threshold (single fruit weight < 20%, shell thickness < 15%, kernel longitudinal diameter < 15%, kernel transverse diameter < 15%, unsaturated fatty acid < 10%, essential amino acid < 10%, palmitoleic acid < 20%, linoleic acid < 30%, oleic acid < 10%, and crude fat < 15%) is screened. The application overcomes the defects of the traditional method of screening only by the fruit setting rate, and integrates the fruit traits into the self-pollination compatibility evaluation system, and can accurately identify the macadamia germplasm that truly has the self-pollination fruit setting ability and excellent quality.

Description

Technical Field

[0001] This invention belongs to the field of plant breeding technology, and in particular relates to a method and application for screening self-compatible and high-quality macadamia germplasm. Background Technology

[0002] Macadamia spp. is a perennial nut crop with high economic value. Its kernels are rich in unsaturated fatty acids, essential amino acids, and other nutrients, giving it a broad market prospect. However, macadamia nuts generally exhibit strong self-incompatibility, resulting in low fruit set rates and large annual yield fluctuations when using single varieties in orchards. This is a core bottleneck restricting the industry's ability to achieve stable and efficient production. Therefore, screening for self-compatible germplasm and cultivating high-quality self-compatible varieties are important research directions in the field of macadamia nut genetics and breeding.

[0003] Currently, the identification and screening of macadamia nut self-compatibility in this field generally uses the self-pollination fruit set rate as the sole evaluation indicator. That is, through bagged self-pollination experiments, the compatibility of germplasm is determined based on the fruit set rate. This method is simple to operate and can achieve rapid initial screening of a large number of germplasms, but it has obvious technical shortcomings.

[0004] First, the evaluation criteria are too simplistic and fail to reflect the actual commercial value of self-pollinated fruits. The self-pollination fruit set rate only reflects the germplasm's ability to set fruit, not the quality of the fruit after setting. Research and production practices have shown that some germplasms with satisfactory self-pollination fruit set rates exhibit significant self-pollination depression in their offspring, manifested as decreased single-fruit weight, thickened shells, and poor kernel development. This directly leads to a reduction in the commercial grade of the fruit and a loss of economic value.

[0005] Secondly, the screening methods neglect the degradation effect on nutritional quality, resulting in a disconnect between the screening results and breeding objectives. Unsaturated fatty acids, essential amino acids, and other components are the core nutritional value of macadamia nuts. However, existing screening methods do not systematically evaluate the intrinsic nutritional quality of fruits after self-pollination. Some germplasms experience a significant decline in core nutritional components after self-pollination. Even if they can set fruit normally, they cannot meet the quality requirements for high-quality commercial fruits and are therefore unsuitable for use as high-quality breeding parents.

[0006] Third, there is a lack of a systematic and quantitative comprehensive screening system, resulting in insufficient screening efficiency and accuracy. Existing technologies have not yet established quantitative evaluation standards for the degree of self-pollination depression in macadamia nuts, making it impossible to accurately distinguish between "low-quality compatible germplasm that can only set fruit" and "superior compatible germplasm that has both fruit-setting ability and high-quality traits." This leads to long breeding screening cycles, a high rate of missing high-quality germplasm, and difficulty in efficiently obtaining core breeding materials that can be directly used.

[0007] In summary, existing self-compatibility germplasm screening techniques are insufficient to simultaneously evaluate both fruit setting ability and fruit quality, failing to meet the practical needs of high-yield and high-quality macadamia nut breeding. There is an urgent need to establish a systematic, quantitative, and reproducible screening method to achieve the synergistic identification of self-compatibility and fruit commercial quality. Summary of the Invention

[0008] In view of this, the purpose of this invention is to provide a method and application for screening self-compatible and high-quality macadamia nut germplasm, which not only evaluates the self-pollination and fruit-setting ability of germplasm, but also comprehensively evaluates the fruit phenotype and nutritional quality of its self-pollinated offspring, thereby achieving efficient and accurate screening of candidate germplasm with both self-compatibility and excellent commercial quality from a large number of germplasm resources.

[0009] To achieve the above-mentioned objectives, the present invention provides the following technical solution: A method for screening self-compatible and high-quality macadamia germplasm includes the following steps: S1: Bagging and self-pollination and natural pollination are carried out on the germplasm to be screened; S2: After the physiological fruit drop period, the self-pollination fruit set rate is counted, and germplasm with a self-pollination fruit set rate ≥ 0.5% is selected and recorded as the initial screening compatible germplasm. S3: After the fruit matures, collect self-pollinated and naturally pollinated fruits of the pre-screened compatible germplasm and determine the phenotypic and nutritional quality traits of the fruit; the phenotypic traits include single fruit weight, shell thickness, kernel longitudinal diameter and kernel transverse diameter; the nutritional quality traits include total unsaturated fatty acids, total essential amino acids, total palmitic acid, total linoleic acid, total oleic acid and total crude fat. S4: Compare the measurement results of self-pollinated fruits and naturally pollinated fruits, and calculate the self-pollination degradation rate; the formula for calculating the self-pollination degradation rate is: Self-pollination depression rate = [(naturally pollinated fruit trait value - self-pollinated fruit trait value) / naturally pollinated fruit trait value] × 100%; S5: Select germplasm with the following characteristics: single fruit weight self-pollination degradation rate <20%, shell thickness self-pollination degradation rate <15%, kernel longitudinal diameter self-pollination degradation rate <15%, kernel transverse diameter self-pollination degradation rate <15%, total unsaturated fatty acid degradation rate <10%, total essential amino acid degradation rate <10%, total palmitic acid self-pollination degradation rate <20%, total linoleic acid self-pollination degradation rate <30%, total oleic acid self-pollination degradation rate <10%, and total crude fat self-pollination degradation rate <15%. These are recorded as self-compatible and high-quality germplasm.

[0010] Preferably, in step S1, bagging self-pollination and natural pollination are carried out during the full flowering period of the germplasm to be screened.

[0011] Preferably, in step S1, each sample of germplasm to be screened selects no fewer than 3 mature trees as biological replicates, and each tree selects no fewer than 100 inflorescences.

[0012] Preferably, in step S5, when the self-growth rate is ≤0, it is considered as no decay.

[0013] Preferably, in S5, the essential amino acids include threonine, valine, methionine, isoleucine, leucine, phenylalanine, and lysine.

[0014] Preferably, it also includes S6: hybridizing the selected self-compatible and high-quality germplasm with the main cultivated variety, and performing two generations of self-pollination verification on the hybrid offspring to ensure that the self-pollination fruit setting rate is ≥0.5% and the self-pollination degradation rate of each fruit trait meets the screening requirements.

[0015] This invention also provides the above-mentioned screening method and the application of the selected self-compatible and high-quality germplasm in macadamia nut breeding.

[0016] Compared with the prior art, the present invention has the following beneficial effects: This invention breaks through the traditional screening model that only uses the self-pollination fruit set rate as a single indicator. On the basis of retaining compatible germplasm in the initial screening of fruit set rate, it further introduces the self-pollination degradation rate to carry out the re-screening of fruit quality. It can accurately remove pseudo-compatible germplasm that "can self-pollinate and bear fruit but whose traits are severely degraded", and greatly reduce the probability of misjudgment and omission of high-quality germplasm.

[0017] The self-compatible germplasm obtained through screening by this invention not only meets the production threshold for self-pollination fruit setting rate, but also controls the self-pollination degradation rate of key commercial traits within a reasonable range: single fruit weight degradation rate is less than 20%, fruit shell thickness degradation rate is less than 15%, total unsaturated fatty acid degradation rate is less than 10%, and total essential amino acid degradation rate is less than 10%. The fruit phenotype and nutritional quality are close to the level of natural cross-pollination, and can be directly used as parent material for the breeding of new self-compatible varieties without the need for additional quality verification, effectively shortening the breeding cycle.

[0018] This invention is the first to systematically quantify the impact of self-inflicted depression on the phenotypic and nutritional quality of macadamia nuts, clarifies the calculation methods and screening thresholds for the self-inflicted depression rate of each core trait, and establishes a standardized screening process. The methods are clearly defined, the indicators are measurable, and the judgment criteria are unified, allowing for reproducible operations by different breeding units, thus facilitating its widespread application in large-scale germplasm resource identification.

[0019] The high-quality self-compatible germplasm obtained through the screening of this invention can provide core gene resources for the improvement of macadamia nut varieties; the self-compatible varieties cultivated can reduce the difficulty of pollinator tree configuration in orchards, weaken the negative impact of adverse weather during flowering on pollination and fertilization, and achieve a dual improvement in orchard yield stability and fruit commercial quality, which has important practical application value for increasing industry efficiency. Attached Figure Description

[0020] Figure 1 This is a flowchart illustrating the method for screening self-compatible and high-quality macadamia germplasm according to the present invention. Detailed Implementation

[0021] This invention provides a method for screening self-compatible and high-quality macadamia germplasm, comprising the following steps: S1. Experimental setup: The germplasm to be screened was subjected to bagging for self-pollination and natural pollination.

[0022] In this invention, preferably during the peak flowering period, bagging self-pollination treatment and natural pollination treatment are carried out simultaneously on the same plant of the germplasm to be screened; further preferably, no less than 3 mature trees are selected for each germplasm as biological replicates, and no less than 100 inflorescences are selected from each tree.

[0023] In this invention, the peak flowering period is further preferred to be the time period during which the flowering volume reaches a peak of 50% or more.

[0024] S2. Initial screening of fruit set rate: After the physiological fruit drop period (about 8 weeks after flowering), the self-pollination fruit set rate is counted, and germplasm with a self-pollination fruit set rate ≥ 0.5% is selected and recorded as the initial screening compatible germplasm.

[0025] The formula for calculating the effective fruit set rate is: .

[0026] S3. Fruit phenotypic and quality determination: After the fruit matures (about 28 weeks after flowering), self-pollinated and naturally pollinated fruits of the pre-screened compatible germplasm are collected, and the phenotypic and nutritional quality traits of the fruit are determined.

[0027] In this invention, the preferred fruit phenotypic traits include kernel longitudinal diameter, transverse diameter, single fruit weight, and shell thickness.

[0028] In this invention, the preferred nutritional quality traits of the fruit include total unsaturated fatty acids, total essential amino acids, total palmitic acid, total linoleic acid, total oleic acid, and total crude fat; further preferred, the nutritional quality traits of the fruit are rapidly and non-destructively determined using near-infrared spectroscopy.

[0029] S4. Self-pollination depression assessment: Compare the measurement results of self-pollinated fruits and naturally pollinated fruits, and calculate the self-pollination depression rate.

[0030] The formula for calculating the auto-decay rate is: .

[0031] S5. Comprehensive screening: Select germplasm with the following characteristics: self-pollination degradation rate of single fruit weight <20%, self-pollination degradation rate of shell thickness <15%, self-pollination degradation rate of kernel longitudinal diameter <15%, self-pollination degradation rate of kernel transverse diameter <15%, degradation rate of total unsaturated fatty acids <10%, degradation rate of total essential amino acids <10%, self-pollination degradation rate of total palmitic acid <20%, self-pollination degradation rate of total linoleic acid <30%, self-pollination degradation rate of total oleic acid <10%, and self-pollination degradation rate of total crude fat <15%. These germplasm are recorded as self-compatible and high-quality germplasm.

[0032] In this invention, it is preferable that when the self-crossing degradation rate of a certain trait is ≤0, it is considered as no degradation.

[0033] In this invention, the preferred essential amino acids include threonine, valine, methionine, isoleucine, leucine, phenylalanine, and lysine.

[0034] S6. Two consecutive generations of self-pollination verification: The selected self-compatible and high-quality germplasm is crossed with the main cultivar, and the hybrid offspring are subjected to two generations of self-pollination verification (F2 and F3 generations) to ensure that the self-pollination fruit set rate is ≥0.5% and the self-pollination degradation rate of each fruit trait meets the screening requirements. As one possible implementation method, the main cultivar is Guire No. 1.

[0035] In this invention, bagging, pollination, and phenotypic trait detection all employ methods commonly used in the field.

[0036] This invention also provides the above-mentioned screening method and the application of the selected self-compatible and high-quality germplasm in macadamia nut breeding.

[0037] The technical solutions provided by the present invention will be described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0038] Example 1 A method for screening self-compatible and high-quality macadamia germplasm, comprising the following steps: S1. Test Setup: Eighty macadamia germplasm resources (from the National Macadamia Germplasm Resource Center, covering commercial varieties, local varieties and wild resources) were selected. During the peak flowering period, three mature trees were selected from each germplasm as biological replicates, and 200 inflorescences were selected from each tree. They were then subjected to bagged self-pollination and tagged natural pollination (control).

[0039] S2, Initial screening of fruit setting rate: After the physiological fruit drop period (approximately 8 weeks after flowering), the number of effective fruits for each treatment was counted, and the self-pollination fruit set rate was calculated.

[0040] The self-pollination fruit set rate of the 80 germplasms ranged from 0.02% to 1.3%, among which 13 germplasms (accounting for 16.3%) had a self-pollination fruit set rate of ≥0.5%, and were recorded as the initial screening compatible germplasms to enter the next stage.

[0041] S3. Fruit phenotypic and quality determination: Thirteen initially screened compatible germplasms were used to harvest self-pollinated and naturally pollinated fruits at the fruit maturity stage (approximately 28 weeks after flowering). Thirty fruits from each sample were randomly selected to determine phenotypic traits (longitudinal and transverse diameter of kernel, single fruit weight, and shell thickness), and nutritional quality (crude fat, fatty acid composition, and amino acid composition) was determined using a near-infrared spectroscopy analyzer.

[0042] S4. Self-fertilization decline assessment: The results showed that the degree of inbreeding degradation varied significantly among different germplasms. HAES814 experienced a 35% decrease in single fruit weight after inbreeding, and NG81 showed a 12% decrease in total unsaturated fatty acids; both were subsequently eliminated.

[0043] S5. Comprehensive Screening: The self-pollination degradation rate of germplasm HAES826 was lower than the threshold in all key traits (single fruit weight degradation rate 8.5%, fruit shell thickness degradation rate -5%, total unsaturated fatty acid degradation rate 2.1%, and total essential amino acid degradation rate 3.5%), and was finally selected as the target germplasm.

[0044] Table 1 shows the comparison data of amino acid composition of self-pollinated and naturally pollinated fruits from some initially screened compatible germplasms. The analysis shows that most non-essential amino acids showed an increasing trend after self-pollination (nutrient concentration effect), which is not a sign of quality decline; the total amount of essential amino acids in HAES826 decreased by only 3.5% after self-pollination, which is far below the 10% threshold and meets the screening criteria.

[0045] Table 1. Amino acid content of self-pollinated and naturally pollinated fruits from different macadamia nut germplasms to be screened.

[0046] Note: In the table, * represents essential amino acids; NP represents natural pollination, and SP represents self-pollination; unit: g / 100g kernels.

[0047] S6. Verification through two consecutive generations of self-crossing: HAES826 was used as the female parent and crossed with the main cultivated variety Guire 1 to obtain the F1 generation. The F1 generation was self-pollinated to produce the F2 generation segregating population (about 500 plants). About 4.4% (22 plants) of the F2 generation had a self-pollution fruit set rate of ≥0.5%, and the selected plants had excellent fruit quality.

[0048] The results of two consecutive generations of self-pollination verification showed that the self-pollination fruit setting rate of F2 and F3 generations was stable at 0.7%~1.1% and 0.6%~0.9%, respectively. The self-pollination degradation rate of key quality traits was consistently below the threshold, confirming its self-compatibility and trait stability.

[0049] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A method for screening self-compatible and high-quality macadamia germplasm, characterized in that, Includes the following steps: S1: Bagging and self-pollination and natural pollination are carried out on the germplasm to be screened; S2: After the physiological fruit drop period, the self-pollination fruit set rate is counted, and germplasm with a self-pollination fruit set rate ≥ 0.5% is selected and recorded as the initial screening compatible germplasm. S3: After the fruit matures, collect self-pollinated and naturally pollinated fruits of the pre-screened compatible germplasm and determine the phenotypic and nutritional quality traits of the fruit; the phenotypic traits include single fruit weight, shell thickness, kernel longitudinal diameter and kernel transverse diameter; the nutritional quality traits include total unsaturated fatty acids, total essential amino acids, total palmitic acid, total linoleic acid, total oleic acid and total crude fat. S4: Compare the measurement results of self-pollinated fruits and naturally pollinated fruits, and calculate the self-pollination degradation rate; the formula for calculating the self-pollination degradation rate is: Self-pollination depression rate = [(naturally pollinated fruit trait value - self-pollinated fruit trait value) / naturally pollinated fruit trait value] × 100%; S5: Select germplasm with the following characteristics: single fruit weight self-pollination degradation rate <20%, shell thickness self-pollination degradation rate <15%, kernel longitudinal diameter self-pollination degradation rate <15%, kernel transverse diameter self-pollination degradation rate <15%, total unsaturated fatty acid degradation rate <10%, total essential amino acid degradation rate <10%, total palmitic acid self-pollination degradation rate <20%, total linoleic acid self-pollination degradation rate <30%, total oleic acid self-pollination degradation rate <10%, and total crude fat self-pollination degradation rate <15%. These are recorded as self-compatible and high-quality germplasm.

2. The screening method according to claim 1, characterized in that, In S1, bagging self-pollination and natural pollination are carried out during the full bloom period of the germplasm to be screened.

3. The screening method according to claim 1, characterized in that, In step S1, each sample of germplasm to be screened shall select no fewer than 3 mature trees as biological replicates, and each tree shall select no fewer than 100 inflorescences.

4. The screening method according to claim 1, characterized in that, In S5, when the self-growth rate is ≤0, it is considered as no decay.

5. The screening method according to claim 1, characterized in that, In S5, the essential amino acids include threonine, valine, methionine, isoleucine, leucine, phenylalanine, and lysine.

6. The screening method according to claim 1, characterized in that, It also includes S6: hybridizing the selected self-compatible and high-quality germplasm with the main cultivated variety, and conducting two generations of self-pollination verification on the hybrid offspring to ensure that the self-pollination fruit setting rate is ≥0.5% and that the self-pollination degradation rate of each fruit trait meets the screening requirements.

7. The application of the screening method according to any one of claims 1 to 6 in macadamia nut breeding.

8. The application of self-compatible and high-quality germplasm selected by the screening method according to any one of claims 1 to 6 in macadamia nut breeding.

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