Propagation and application of new apple germplasm obtained through grafting crossbreeding

Abstract

The invention discloses propagation and application of a new apple germplasm obtained through grafting crossbreeding. The invention provides a method for preparing apple fruits, which comprises the following steps of: preparing grafted seedlings by using scions from apples, and harvesting fruits on branches grown from the scions of the grafted seedlings. The invention also provides an apple propagation method, which comprises the following steps: grafting the branch from the apple as a scion to a stock to obtain a grafted seedling. The apple is an apple (Malus pumila) Cui Xiang, and the preservation registration number of the apple is CGMCC (China General Microbiological Culture Collection Center) No. 46277. The inventor of the invention utilizes' Hongyu 'and' Meile 'apple grafted seedlings with strong flavor, and combines an adventitious bud induction technology to realize gene penetration and recombination among grafted bodies, and obtains a new apple variety with strong fruit fragrance, late maturing, storage resistance, early fruit property, yielding ability and disease resistance.

Description

Technical Field

[0001] The invention belongs to the field of biotechnology and relates to the propagation and application of new apple germplasm obtained through grafting and cross breeding. Background Art

[0002] It is a major national need to establish a big food concept, ensure a stable and safe supply of important agricultural products, hold the Chinese fruit plate firmly, cultivate high-quality, late-maturing, storable industry main varieties and characteristic diversified varieties of important deciduous fruit trees, establish an internal circulation of self-bred, self-produced, and self-sold high-quality fruits that does not rely on imports, meet the year-round supply needs of 1.4 billion Chinese people for high-quality fruits, contribute to a healthy China, promote continuous increase in farmers' income, and ensure ecological security.

[0003] Apple is the largest species of deciduous fruit trees in my country, with an area and output accounting for more than 50% of the world's total, and an annual total output value of more than 200 billion yuan. It is a pillar industry for increasing farmers' income and a major fruit product that guarantees the year-round supply of high-quality fruits. However, more than 70% of my country's apple industry is currently Red Fuji bud mutation varieties, which have the following three problems: ① The problems of singleness and homogeneity are prominent, affecting the continuous increase in fruit farmers' income; ② The late-maturing and storage-resistant characteristics of Red Fuji apple varieties are very prominent, but they are mainly sweet and not fragrant; ③ Red Fuji apples are difficult to flower and bear fruit late, and are relatively difficult to manage, which is not conducive to the development of large-scale modern cultivation models. Therefore, in order to optimize and adjust the variety structure, transform and upgrade, and develop high-quality development of my country's apple industry, it is necessary to further select and breed new apple varieties with strong fruit fragrance, late maturation, storage resistance, easy flowering, early fruiting and high yield, and easy management.

[0004] Choosing the right breeding approach is the key to achieving breeding goals. At present, the conventional breeding approaches for fruit trees mainly include hybrid breeding and bud mutation selection. Hybrid breeding is to determine breeding goals based on the industry's variety needs, select and match parents, and select new varieties with excellent comprehensive traits, in line with breeding goals, and strong hybrid vigor through sexual hybridization and the separation, combination, interaction and modification of its genes. Almost all traits can be improved in one hybridization. Bud mutation selection (natural mutation) is aimed at the problems of individual traits of the main cultivated varieties in production, and uses somatic natural mutations and their epigenetics to continuously and repeatedly improve individual traits to meet the needs of production and consumption.

[0005] The current breeding goal is to retain the late-ripening and storage-resistant characteristics of Red Fuji apple varieties while improving some of their characteristics, such as weak fragrance, difficulty in flowering, and late fruiting. Therefore, research and development of new breeding approaches are of great significance to achieving breeding goals. Summary of the invention

[0006] The purpose of the present invention is to provide the propagation and application of new apple germplasm obtained by grafting hybrid breeding.

[0007] The present invention provides a method for preparing apple fruit, comprising the following steps: preparing grafted seedlings using scions from apples, and harvesting fruits from branches grown from the scions of the grafted seedlings; It is characterized in that: the apple is apple ( Malus pumila ) Cui Xiang, whose deposit registration number is CGMCC No.46277.

[0008] Specifically, the preparation method of the grafted seedlings is: grafting a branch segment from apple as a scion onto a rootstock.

[0009] Specifically, the branch segment is a branch segment cut from a branch of the current year, with a length of 8-12 cm and 2-3 buds.

[0010] Specifically, the rootstock is Malus octagonalis.

[0011] Specifically, the method for preparing the rootstock is as follows: a transverse cut is made on the trunk of the Malus octagonalis seedling, and the height of the trunk is retained at 5-10 cm.

[0012] Specifically, the grafting method is branch grafting.

[0013] Specifically, the grafting method is: making a longitudinal incision at the top of the rootstock, grafting the scion in a branch grafting manner, and grafting one scion per plant.

[0014] The present invention also provides an apple propagation method, comprising the following steps: Grafting apple branches as scions onto rootstocks to obtain grafted seedlings; It is characterized in that: the apple is apple ( Malus pumila ) Cui Xiang, whose deposit registration number is CGMCC No.46277.

[0015] Specifically, the branch segment is a branch segment cut from a branch of the current year, with a length of 8-12 cm and 2-3 buds.

[0016] Specifically, the rootstock is Malus octagonalis.

[0017] Specifically, the method for preparing the rootstock is as follows: a transverse cut is made on the trunk of the Malus octagonalis seedling, and the height of the trunk is retained at 5-10 cm.

[0018] Specifically, the grafting method is branch grafting.

[0019] Specifically, the grafting method is: making a longitudinal incision at the top of the rootstock, grafting the scion in a branch grafting manner, and grafting one scion per plant.

[0020] The present invention also provides an apple breeding method, comprising the following steps: (1) preparing a grafted seedling; the main trunk of the grafted seedling comprises, from bottom to top, a rootstock, an interstock and a scion; the interstock is apple variety A; and the scion is apple variety B; (2) Cultivating grafted seedlings and inducing adventitious buds during the cultivation process; (3) Screening variant branches from branches grown from adventitious buds; (4) The variant branches are grafted to obtain new apple germplasm.

[0021] Specifically, the apple variety A is 'Ruby' apple, and the apple variety B is 'Merlot' apple.

[0022] Specifically, in the apple breeding method, the rootstock is Pingyi sweet tea.

[0023] Specifically, in the apple breeding method, the method for preparing grafted seedlings comprises the following steps: In the first year, the trunk of the plant used for making the rootstock was transversely cut, and the trunk was kept at a height of 5-10 cm as the rootstock; a longitudinal incision was made at the top of the rootstock, and the scion from apple variety A was grafted by branch grafting, with one scion grafted per plant, and normal cultivation and management were carried out; In the second year, a transverse excision was made on the main trunk section of apple variety A of the plant, and a 30-40 cm height of the main trunk section of apple variety A was retained as the middle rootstock; a longitudinal incision was made at the top of the middle rootstock, and the scion from apple variety B was grafted by branch grafting, one scion per plant, and normal cultivation and management were carried out.

[0024] Specifically, the scion is a branch segment with a length of 8-12 cm and 2-3 buds cut from the current year's branches.

[0025] The step (2) may specifically include the following steps: In the third year, transverse excision was performed on the trunk section of the grafted apple variety B seedlings to make the trunk height of the grafted seedlings 80-120 cm. Before budding, the main buds and side buds of the Merlot trunk section were removed to induce adventitious buds.

[0026] In the fourth year, the side branches of the main trunk section of apple variety B were removed before the grafted seedlings sprouted, and the main buds and side buds of the main trunk section of apple variety B were removed before sprouting to induce adventitious buds.

[0027] apple( Malus pumila ) Cui Xiang, was deposited on December 6, 2024 in the General Microbiology Center of China Microorganism Culture Collection Committee (referred to as CGMCC, address: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences), and the deposit registration number is CGMCC No.46277.

[0028] The apple fruit has characteristics (a) and / or (b) and / or (c) and / or (d) and / or (e) and / or (f) and / or (g) and / or (h) and / or (i) and / or (j).

[0029] The apple ( Malus pumila ) The apple fruits grown in Cui Xiang have characteristics (a) and / or (b) and / or (c) and / or (d) and / or (e) and / or (f) and / or (g) and / or (h) and / or (i) and / or (j).

[0030] The apple fruit has characteristics (a) and / or (b) and / or (d) and / or (e) and / or (f) and / or (g) and / or (h) and / or (i) and / or (j).

[0031] The apple ( Malus pumila ) The apple fruits grown in Cui Xiang have characteristics (a) and / or (b) and / or (d) and / or (e) and / or (f) and / or (g) and / or (h) and / or (i) and / or (j).

[0032] The apple fruit has characteristics (a) and / or (b) and / or (d) and / or (e) and / or (f) and / or (g) and / or (i) and / or (j).

[0033] The apple ( Malus pumila ) The apple fruits grown in Cui Xiang have characteristics (a) and / or (b) and / or (d) and / or (e) and / or (f) and / or (g) and / or (i) and / or (j).

[0034] The apple fruit has (a) and / or (b) and / or (d) and / or (e) and / or (f) and / or (g).

[0035] The apple ( Malus pumila ) The apple fruits grown in Cui Xiang have (a) and / or (b) and / or (d) and / or (e) and / or (f) and / or (g).

[0036] The apple fruit has (a) and / or (b) and / or (d) and / or (e) and / or (f).

[0037] The apple ( Malus pumila ) The apple fruits grown in Cui Xiang have (a) and / or (b) and / or (d) and / or (e) and / or (f).

[0038] (a) The hardness of apple fruit is 7 kg / cm 2 Specifically, the hardness of apple fruit is 6.5kg / cm2 Specifically, the hardness of apple fruit is 6kg / cm 2 Specifically, the hardness of apple fruit is 5.5 kg / cm 2 the following.

[0039] (b) The sucrose content of the apple fruit is 16µg / g or more. Specifically, the sucrose content of the apple fruit is 17µg / g or more. Specifically, the sucrose content of the apple fruit is 18µg / g or more. Specifically, the sucrose content of the apple fruit is 19µg / g or more. Specifically, the sucrose content of the apple fruit is 20µg / g or more. Specifically, the sucrose content of the apple fruit is 21µg / g or more.

[0040] (c) The sorbitol content of apple fruit is 1100 µg / g or more.

[0041] (d) The galactose content of the apple fruit is 25µg / g or more. Specifically, the galactose content of the apple fruit is 30µg / g or more. Specifically, the galactose content of the apple fruit is 35µg / g or more. Specifically, the galactose content of the apple fruit is 40µg / g or more. Specifically, the galactose content of the apple fruit is 45µg / g or more. Specifically, the galactose content of the apple fruit is 50µg / g or more. Specifically, the galactose content of the apple fruit is 55µg / g or more. Specifically, the galactose content of the apple fruit is 60µg / g or more. Specifically, the galactose content of the apple fruit is 65µg / g or more.

[0042] (e) The trehalose content of the apple fruit is 26 µg / g or more. Specifically, the trehalose content of the apple fruit is 30 µg / g or more. Specifically, the trehalose content of the apple fruit is 35 µg / g or more. Specifically, the trehalose content of the apple fruit is 40 µg / g or more.

[0043] (f) The malic acid content of the apple fruit is 6000µg / g or less. Specifically, the malic acid content of the apple fruit is 5500µg / g or less. Specifically, the malic acid content of the apple fruit is 5000µg / g or less. Specifically, the malic acid content of the apple fruit is 4500µg / g or less. Specifically, the malic acid content of the apple fruit is 4000µg / g or less.

[0044] (g) The aroma substances of apple fruit contain the following 11 substances: 5-heptene-2-ol; 1-tridecan-4-ol; 1-cyclohexyl-2-nitropropane-1,3-diol; n-propyl acetate; 3-methyl-3-butene-1-ol acetate; 2-methylpropionic acid butyl ester; 3-hexen-1-ol acetate; 2-methylbutyric acid pentyl ester; hexyl butyrate; 2-methylbutyric acid hexyl ester; butyl trans-3-enyl phthalate.

[0045] (h) The content of alcohol substances in the aroma substances of apple fruit is 0.013 μg / g or more. Specifically, the content of alcohol substances in the aroma substances of apple fruit is 0.0133 μg / g or more.

[0046] (i) The content of lipid substances in the aroma substances of apple fruits is 0.1µg / g or more. Specifically, the content of lipid substances in the aroma substances of apple fruits is 0.15µg / g or more. Specifically, the content of lipid substances in the aroma substances of apple fruits is 0.2µg / g or more. Specifically, the content of lipid substances in the aroma substances of apple fruits is 0.25µg / g or more. Specifically, the content of lipid substances in the aroma substances of apple fruits is 0.26µg / g or more. Specifically, the content of lipid substances in the aroma substances of apple fruits is 0.27µg / g or more. Specifically, the content of lipid substances in the aroma substances of apple fruits is 0.28µg / g or more. Specifically, the content of lipid substances in the aroma substances of apple fruits is 0.29µg / g or more.

[0047] (j) The content of aldehydes in the aroma substances of apple fruits is 0.4µg / g or more. Specifically, the content of aldehydes in the aroma substances of apple fruits is 0.45µg / g or more. Specifically, the content of aldehydes in the aroma substances of apple fruits is 0.46µg / g or more. Specifically, the content of aldehydes in the aroma substances of apple fruits is 0.47µg / g or more. Specifically, the content of aldehydes in the aroma substances of apple fruits is 0.48µg / g or more. Specifically, the content of aldehydes in the aroma substances of apple fruits is 0.49µg / g or more. Specifically, the content of aldehydes in the aroma substances of apple fruits is 0.50µg / g or more. Specifically, the content of aldehydes in the aroma substances of apple fruits is 0.51µg / g or more.

[0048] µg / g, the denominator g refers to the fresh weight of apple fruit.

[0049] When apple variety B is grafted onto apple variety A, genetic materials such as DNA and RNA of apple variety A can penetrate into apple variety B through the vascular system, and genetic recombination occurs, leading to trait variation.

[0050] The inventors of the present invention have aimed at the problems of Red Fuji apple varieties such as weak fruit fragrance, difficulty in flowering, late fruiting, and difficulty in management. Based on the technical principles of asexual hybridization breeding of fruit trees, the inventors of the present invention used grafted seedlings of 'Red Jade' and 'Merlot' apples with strong flavor, combined with adventitious bud induction technology, to achieve gene penetration and recombination between grafted bodies, and obtained a new apple variety with strong fruit fragrance, late maturity, storability, early fruiting, high yield and strong disease resistance. BRIEF DESCRIPTION OF THE DRAWINGS

[0051] Figure 1 These are fruits picked from the plants in the test group in Example 3 without bagging (illustrative photo of a single fruit).

[0052] Figure 2 These are fruits picked after bagging treatment of the test group plants in Example 3 (illustrative photos of single fruits).

[0053] Figure 3 These are fruits picked from the control group plants after bagging treatment in Example 3 (illustrative photos of single fruits).

[0054] Figure 4 These are fruits picked after the plants in the test group in Example 3 were bagged (illustrative photo of a group of fruits).

[0055] Figure 5 This is a partially enlarged photo of the fruits picked after the plants in the test group in Example 3 were bagged.

[0056] Figure 6 This is a partially enlarged photograph of the fruits picked after the plants in the control group were bagged in Example 3.

[0057] Figure 7 These are the results of ISSR detection of Wanglin, Meiluo, Hongyu and Cuixiang in Example 4.

[0058] Figure 8 This is the evolutionary tree of Wanglin, Meiluo, Hongyu and Cuixiang in Example 4.

[0059] Fig. 9 Schematic diagram of grafting. DETAILED DESCRIPTION

[0060] The present invention is further described in detail below in conjunction with specific embodiments, and the examples provided are only for illustrating the present invention, rather than for limiting the scope of the present invention. The examples provided below can be used as a guide for further improvements by those of ordinary skill in the art, and do not constitute a limitation of the present invention in any way.

[0061] The experimental methods in the following embodiments, unless otherwise specified, are conventional methods, and are carried out in accordance with the techniques or conditions described in the literature in the field or in accordance with the product instructions. The materials, reagents, etc. used in the following embodiments, unless otherwise specified, can be obtained from commercial sources. Unless otherwise specified, the quantitative tests in the following embodiments are set up with three repeated experiments, and the results are averaged. In the embodiments, the hardness of apple fruit is detected by a texture analyzer. In the embodiments, the sugar content and acid content of apple fruit are detected by ultra-performance liquid chromatography tandem triple quadrupole mass spectrometry (UPLC-MS / MS). In the embodiments, the aroma substance content of apple fruit is detected by headspace solid phase microextraction (HS-SPME). See the schematic diagram of grafting. Fig. 9 .

[0062] The 'Ruby' apple is native to the United States. It was first discovered in 1800 in LJlster County, New York (known as Jonathan in the United States). It was widely developed from the late 19th century to the early 20th century and cultivated in all continents of the world. It was introduced to the Liaodong Peninsula in China in the early 20th century.

[0063] The parents of Fuji apples are Guoguang and Yuanshuai, which were hybridized in 1939, named and registered in 1962, and introduced to my country in 1965. The Fuji apples first cultivated in Japan had poor coloring. In the subsequent promotion and cultivation process, Japanese scientific research and promotion institutions gradually selected many colored Fuji bud mutations from Fuji, collectively known as colored Fuji (or simply "Red Fuji"). "Nagafu No. 2" is a colored Fuji variety selected from Nagano Prefecture, Japan. It was declared in 1975 and approved in 1981. The Fruit Tree Branch of Yantai Academy of Agricultural Sciences selected a new Fuji dark red bud mutation variety "Merlot" from "Nagafu No. 2". "Merlot" apples have outstanding characteristics such as good fruit coloring and fast coloring. In 2014, it passed the new variety approval of Shandong Province. 'Merlot' apple is recorded in the following literature: References: Song Laiqing, Zhao Lingling, et al., Selection of Red Fuji apple bud mutations and breeding of new varieties, Yantai Fruit Trees 2015, 4: 6-7.

[0064] Example 1: Establishing a new apple breeding method and obtaining excellent new germplasm The inventors have established a new apple breeding method: using grafting and hybridization breeding to obtain new apple germplasm.

[0065] 1. Preparation of grafted seedlings ①In March 2013, Pingyi sweet tea seedlings with a height of 40-50 cm were planted in a nursery in Yantai City, Shandong Province and cultivated and managed normally.

[0066] ②In March 2014, the trunk of the plant in step ① was transversely cut, and the trunk was kept at a height of 5-10 cm as the rootstock. A longitudinal incision was made at the top of the rootstock, and the scion was grafted by branch grafting (the preparation method of the scion: take the current year's branches of the 'Ruby' apple plant, cut a branch segment with a length of about 10 cm and 2-3 buds), and each plant was grafted with one scion and cultivated and managed normally.

[0067] ③In March 2015, the ruby ​​trunk section of the plant in step ② was transversely cut, and the ruby ​​trunk section was retained at a height of 30-40 cm as the middle stock. A longitudinal incision was made at the top of the middle stock, and the scion was grafted by branch grafting (the preparation method of the scion: take the current year branches of the 'Merlot' apple plant, cut a branch section with a length of about 10 cm and 2-3 buds), and each plant was grafted with one scion and cultivated and managed normally.

[0068] According to the above steps, 40 grafted seedlings were obtained. The main trunk of the grafted seedlings was composed of a 5-10 cm high rootstock (Pingyi sweet tea), a 30-40 cm high intermediate rootstock ('Hongyu' apple), and a scion ('Melody' apple).

[0069] 2. In March 2016, the main trunk section of the grafted seedlings in step 1 was transversely cut to make the trunk height of the grafted seedlings 80-120cm, and transplanted to the apple orchard (with a plant spacing of 2m and a row spacing of 5m). The main buds and side buds of the main trunk section of the Merlot were removed before budding to induce adventitious buds. Normal cultivation and management.

[0070] 3. In March 2017, the side branches of the main trunk section of the Merlot were removed before the grafted seedlings sprouted in step 2, and the main buds and side buds of the main trunk section of the Merlot were removed before sprouting to induce adventitious buds. Normal cultivation and management.

[0071] 4. In September 2019, a variant branch was found in the main trunk section of a Merlot plant grafted in step 3 (the variant branch is a branch growing on the main trunk, also known as the main branch variant branch). The phenotype of the apple fruits on the variant branch is: the skin is green, the fruit surface is smooth, and there are almost no star spots; the flesh is crispy, sweet, and has a strong aroma. The phenotype of the fruits on the branches other than the variant branch on the grafted seedling and on each branch on other grafted seedlings is: the skin is red, the fruit surface has many star spots, and it is relatively rough; the crispness, sweetness, and aroma of the flesh are not as good as those of the apple fruits on the variant branch.

[0072] Example 2: Propagation of excellent new germplasm and evaluation of trait stability 1. In March 2019, Malus multiflora seedlings with a height of 40-50 cm (with a plant spacing of 2 m and a row spacing of 4 m) were planted in an apple orchard in Yantai City, Shandong Province, and normal cultivation and management were carried out.

[0073] 2. In March 2020, transverse incision was made on the trunk of the plant in step 1, and the trunk was retained at a height of 5-10 cm as the rootstock. Then 300 grafted seedlings of the experimental group and 80 grafted seedlings of the control group were prepared respectively. Preparation method of the grafted seedlings in the experimental group: a longitudinal incision was made at the top of the rootstock, and the scion was grafted by branch grafting (the preparation method of the scion: a branch segment with a length of about 10 cm and 2-3 buds was cut from the current year branch on the variant branch in step 4 of Example 1), and one scion was grafted for each plant. Preparation method of the grafted seedlings in the control group: a longitudinal incision was made at the top of the rootstock, and the scion was grafted by branch grafting (the preparation method of the scion: a branch segment with a length of about 10 cm and 2-3 buds was cut from the current year branch of the 'Merlot' apple plant), and one scion was grafted for each plant. Normal cultivation and management.

[0074] 3. Characteristics in 2022.

[0075] From April to May, the flowering was observed. The experimental group plants had a large number of flowers on their branches, with each plant having 50-80 flowers. The control group plants had sporadic flowers on their branches, with each plant having 10-20 flowers.

[0076] The apple fruits of both the experimental group and the control group matured in late October and are late-maturing varieties. The number of apple fruits on the experimental group plants was significantly higher than that on the control group plants. The number of apple fruits on each plant in the experimental group plants was 30-50. The number of apple fruits on each plant in the control group plants was 5-10. The characteristics of the apple fruits on the experimental group plants: the skin is green, the fruit surface is smooth, and there are almost no star spots; the flesh is crispy, sweet, and has a strong aroma. The characteristics of the apple fruits on the control group plants: the skin is red, the fruit surface has many star spots, and it is relatively rough; the crispness, sweetness, and aroma of the flesh are not as good as those of the apple fruits on the experimental group plants.

[0077] The apple fruits of the experimental group and the control group were sampled respectively to detect the hardness, sugar content (sucrose content, sorbitol content, galactose content and trehalose content), acid content (malic acid content) and aroma substance content.

[0078] 4. Characteristics in 2023.

[0079] From April to May, the flowering situation was observed. The number of flowers in the experimental group was significantly higher than that in the control group. The number of flowers in the experimental group was 150-200 per plant. The number of flowers in the control group was 50-100 per plant.

[0080] The apple fruits of both the experimental group and the control group matured in late October and were late-maturing varieties. The number of apple fruits in the experimental group was significantly higher than that in the control group. The number of apple fruits per plant in the experimental group was 100-150. The number of apple fruits per plant in the control group was 40-80. The characteristics of the apple fruits on the experimental group plants: the peel was green, the fruit surface was smooth, and there were almost no star spots; the flesh was crispy and sweet, with a strong aroma. The characteristics of the apple fruits on the control group plants: the peel was red, the fruit surface had many star spots, and it was rough; the crispness, sweetness and aroma of the flesh were not as good as those of the apple fruits on the experimental group plants. The average single fruit weight of the apple fruits of the experimental group plants was 316.7g±33.70g. The average single fruit weight of the apple fruits of the control group plants was 286.2g±21.58g.

[0081] The apple fruits of the experimental group and the control group were sampled respectively to detect the hardness, sugar content (sucrose content, sorbitol content, galactose content and trehalose content), acid content (malic acid content) and aroma substance content.

[0082] There was no pre-harvest fruit drop in the experimental group and the control group. There was no branch ring disease phenotype in the experimental group and no early leaf fall phenotype. There was no early leaf fall phenotype in the control group, and 5% (4 / 80) had branch ring disease phenotype.

[0083] 5. Characteristics in 2024.

[0084] From April to May, the flowering situation was observed. The number of flowers in the experimental group was significantly higher than that in the control group. The number of flowers in the experimental group was 600-800 per plant. The number of flowers in the control group was 200-300 per plant.

[0085] The apple fruits of both the experimental group and the control group matured in late October and were late-maturing varieties. The number of apple fruits in the experimental group was significantly higher than that in the control group. The number of apple fruits per plant in the experimental group was 160-200. The number of apple fruits per plant in the control group was 80-140. The characteristics of the apple fruits on the experimental group plants: the peel was green, the fruit surface was smooth, and there were almost no star spots; the flesh was crispy and sweet, with a strong aroma. The characteristics of the apple fruits on the control group plants: the peel was red, the fruit surface had many star spots, and it was rough; the crispness, sweetness and aroma of the flesh were not as good as those of the apple fruits on the experimental group plants. The average single fruit weight of the apple fruits of the experimental group plants was 324.6g±28.64g. The average single fruit weight of the apple fruits of the control group plants was 285.9g±22.18g.

[0086] The apple fruits of the experimental group and the control group were sampled respectively to detect the hardness, sugar content (sucrose content, sorbitol content, galactose content and trehalose content), acid content (malic acid content) and aroma substance content.

[0087] There was no pre-harvest fruit drop in the experimental group and the control group. There was no branch ring disease phenotype in the experimental group and no early leaf fall phenotype. There was no early leaf fall phenotype in the control group, and 6.25% (5 / 80) of the plants had branch ring disease phenotype.

[0088] 6. Result data The results of the firmness of apple fruits in 2022, 2023, and 2024 are shown in Table 1 (sample size N=50). The apple fruits in the experimental group were less firm than those in the control group and therefore more crispy.

[0089] Table 1

[0090] The results of sugar content and acid content of apple fruits in 2022, 2023 and 2024 are shown in Table 2 (the units are all µg / g, and the denominator g refers to the fresh weight of the apple fruit) (sample number N=50). The sucrose content, sorbitol content, galactose content and trehalose content of apple fruits in the experimental group were significantly higher than those in the control group, and the malic acid content of apple fruits in the experimental group was significantly lower than that in the control group. Therefore, compared with the apple fruits in the control group, the apple fruits in the experimental group were sweeter and less acidic.

[0091] Table 2

[0092] In 2024, the results of the aroma substance content of apple fruits in the control group are shown in Table 3, and the results of the aroma substance content of apple fruits in the experimental group are shown in Table 4. The units are all μg / g (the denominator g refers to the fresh weight of apple fruits), and the number of samples N=50. The results show that the content of alcohol substances, ester substance yield and aldehyde substance content in apple fruits in the experimental group are significantly higher than those in apples in the control group. Compared with apple fruits in the control group, there are 11 specific aroma substances in apples in the experimental group: 5-heptene-2-ol; 1-tridecan-4-ol; 1-cyclohexyl-2-nitropropane-1,3-diol; n-propyl acetate; 3-methyl-3-butene-1-ol acetate; 2-methylpropionic acid butyl ester; 3-hexen-1-ol acetate; 2-methylbutyric acid pentyl ester; butyric acid hexyl ester; 2-methylbutyric acid hexyl ester; butyl trans-3-enyl phthalate. The data of 2022 and 2023 are consistent with the data of 2024.

[0093] Table 3

[0094] Table 4

[0095] The above results show that a new apple germplasm with strong fragrance, easy flowering, early fruit setting and high fruit setting rate has been obtained. In addition, the new apple germplasm has strong disease resistance (no branch ring rot and early leaf fall diseases), no pre-harvest fruit drop, late maturity, storage resistance and good management. After three consecutive years of observation, the new apple germplasm has stable characteristics. The new apple germplasm was named Apple ( Malus pumila ) Cui Xiang, also known as apple 'Cui Xiang'.

[0096] apple( Malus pumila ) Cui Xiang, was deposited on December 6, 2024 in the General Microbiology Center of China Microorganism Culture Collection Committee (referred to as CGMCC, address: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences), and the deposit registration number is CGMCC No.46277.

[0097] Example 3: Color properties of apple 'Cui Xiang' 1. In March 2020, Malus multiflora seedlings with a height of 40-50 cm (with a plant spacing of 2 m and a row spacing of 4 m) were planted in an orchard in Yantai City, Shandong Province, and normal cultivation and management were carried out.

[0098] 2. In March 2021, make a transverse cut on the trunk of the plant in step 1, and keep 5-10 cm of the trunk as the rootstock. Then prepare 100 grafted seedlings for the test group and 80 grafted seedlings for the control group. Normal cultivation and management.

[0099] Preparation method of the grafted seedlings in the test group: make a longitudinal incision at the top of the rootstock and graft the scion by branch grafting (preparation method of the scion: take the current year branches of the grafted seedlings in the test group of Example 2, cut a branch segment with a length of about 10 cm and 2-3 buds), and graft one scion per plant.

[0100] Preparation method of the grafted seedlings in the control group: make a longitudinal incision at the top of the rootstock and graft the scion by branch grafting (preparation method of the scion: take the current year branches of the grafted seedlings in the control group of Example 2, cut the branch segments with a length of about 10 cm and 2-3 buds), and graft one scion per plant.

[0101] 3. In 2024, some fruits will be bagged 30 days after the flowers fall (the bags will be removed after the fruits are ripe in October and picked 10 days after the bags are removed), and some fruits will not be bagged and will be cultivated and managed normally.

[0102] The fruit of the experimental group plants after picking without bagging (see the exemplary photo) Figure 1 ): The fruit skin is green and the surface is smooth. Fruits picked from the experimental group after being bagged (see the photo of a single fruit for an example) Figure 2 , see the sample photos of the group fruit Figure 4 , see the enlarged photo of the part Figure 5 ): The base color is light yellow, and some of the fruit surface is pink, which is very beautiful. The fruit of the control group plants that were not bagged after picking: the skin is bright red. The fruit of the control group plants that were bagged after picking (see the exemplary photo) Figure 3 , see the enlarged photo of the part Figure 6 ): The peel is dark red.

[0103] Example 4. Genomic and DNA fingerprint (ISSR) detection and analysis of variant sources Test samples: Leaves of apple 'Cui Xiang', apple 'Mei Le', apple 'Hong Yu' or apple 'Wang Lin'. Take the test samples, extract genomic DNA, perform ISSR detection and genome sequencing, and construct a phylogenetic tree. The results of ISSR detection of Mei Le, Hong Yu and Cui Xiang are shown in Figure 7 . Evolutionary tree see Figure 8 .

[0104] The results showed that there were significant differences in the genomes of the tested samples. The variation of Apple'Cui Xiang' was a variation of genetic material, mainly the penetration and recombination of genetic materials such as DNA and RNA between Apple'Meile' and Apple'Hongyu'. In the evolutionary tree, Apple'Cui Xiang' and Apple'Meile' were first clustered into one category, and then clustered into one category with Apple'Hongyu', while Apple'Wanglin' ​​was clustered into a separate category, indicating that Apple'Cui Xiang' was more closely related to Apple'Meile' and Apple'Hongyu', and more distantly related to Apple'Wanglin'.

[0105] The present invention has been described in detail above. It will be apparent to those skilled in the art that the present invention may be implemented in a wide range under equivalent parameters, concentrations and conditions without departing from the spirit and scope of the present invention and without the need for unnecessary experimentation. Although the present invention provides specific embodiments, it should be understood that further improvements may be made to the present invention. In short, according to the principles of the present invention, this application intends to include any changes, uses or improvements to the present invention, including changes made by conventional techniques known in the art that depart from the scope disclosed in this application. Applications of some of the basic features may be made within the scope of the following appended claims.

Claims

1. A method for preparing apple fruit, comprising the following steps: preparing grafted seedlings using scions from apples, and harvesting fruits from branches grown from the scions of the grafted seedlings; Features: The apple is apple ( Malus pumila ) Cui Xiang, whose deposit registration number is CGMCC No.46277.

2. The method according to claim 1, characterized in that: The preparation method of the grafted seedling is as follows: a branch segment from apple is used as a scion and grafted onto a rootstock.

3. The method according to claim 2, characterized in that: The branch segments are 8-12 cm long and have 2-3 buds, cut from current year branches.

4. The method according to claim 2 or 3, characterized in that: The rootstock is Malus octagonalis.

5. A method for propagating apples, comprising the following steps: Grafting apple branches as scions onto rootstocks to obtain grafted seedlings; Features: The apple is apple ( Malus pumila ) Cui Xiang, whose deposit registration number is CGMCC No.46277.

6. The method according to claim 5, characterized in that: The branch segments are 8-12 cm long and have 2-3 buds, cut from current year branches.

7. The method according to claim 5 or 6, characterized in that: The rootstock is Malus octagonalis.

8. An apple breeding method comprising the following steps: (1) preparing a grafted seedling; the main trunk of the grafted seedling comprises, from bottom to top, a rootstock, an interstock and a scion; the interstock is apple variety A; and the scion is apple variety B; (2) Cultivating grafted seedlings and inducing adventitious buds during the cultivation process; (3) Screening variant branches from branches grown from adventitious buds; (4) The variant branches are grafted to obtain new apple germplasm.

9. The method according to claim 8, characterized in that: The apple variety A is 'Ruby' apple, and the apple variety B is 'Merlot' apple.

Images