CN115997676A - Breeding method of white loquat and method for shortening childhood - Google Patents
Breeding method of white loquat and method for shortening childhood Download PDFInfo
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Classifications
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H1/00—Processes for modifying genotypes ; Plants characterised by associated natural traits
- A01H1/02—Methods or apparatus for hybridisation; Artificial pollination ; Fertility
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H5/00—Angiosperms, i.e. flowering plants, characterised by their plant parts; Angiosperms characterised otherwise than by their botanic taxonomy
- A01H5/08—Fruits
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H6/00—Angiosperms, i.e. flowering plants, characterised by their botanic taxonomy
- A01H6/74—Rosaceae, e.g. strawberry, apple, almonds, pear, rose, blackberries or raspberries
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- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Botany (AREA)
- Developmental Biology & Embryology (AREA)
- Environmental Sciences (AREA)
- Physiology (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Natural Medicines & Medicinal Plants (AREA)
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
- Pretreatment Of Seeds And Plants (AREA)
Abstract
The invention provides a breeding method of white loquat and a method for shortening childhood, and relates to the technical field of agricultural breeding. The invention relates to a breeding method of white loquat and a method for shortening childhood, which concretely comprise the following steps: s1 parent selection, S2 pollen preparation, S3 hybridization assembly, S4 cold protection and fruit preservation, S5 sowing and seedling cultivation, S6 nursery establishment, S7 transplanting and field planting, S8 sapling fertilization, S9 sapling shaping, S10 comprehensive childhood shortening technology and the like. The method improves the screening efficiency of the hybrid parents and the hybrid offspring of the white-meat loquat, improves the preservation time and pollination efficiency of the loquat pollen, improves the yield of hybrid seeds, avoids the influence of grafting or repeated application of high-concentration hormone on the plant characters of the hybrid offspring of the loquat, shortens the childhood period of the hybrid offspring of the loquat by more than 1 year, and can lead the flowering rate of the hybrid offspring of the loquat after sowing to be more than 77.5 percent at the 4 th year.
Description
Technical Field
The invention belongs to the technical field of agricultural breeding, and particularly relates to a breeding method of white-meat loquat and a method for shortening childhood.
Background
Loquat is a famous and distinctive tree species of original China, and has extremely rich resources. The loquat variety can be classified into white loquat variety and yellow loquat variety according to the pulp color. The pulp of the white-meat loquat variety is milky or light yellow, the pulp is tender, the juice is sweet, the flavor is excellent, the quality is better than that of the yellow-meat variety, and the white-meat loquat variety is most suitable for fresh food. Although the quality of the fruits of the white loquat variety is better than that of the yellow loquat variety, the main variety cultivated in the world is the yellow loquat variety, and the white loquat variety mainly occupies a certain proportion in Jiangsu, zhejiang, shanghai and other places in northern loquat producing areas in China, and the trace thereof is difficult to see in other loquat producing areas. An important reason for this is the small number of white meat varieties, and the inability of each production zone to find a suitable good variety from a limited white meat variety source. Therefore, the mass cultivation of the fine white loquat variety is urgent for adjusting the variety structure of the loquat industry in China and even worldwide.
But the innovation of the breeding method is key to cultivate a great deal of fine white loquat varieties. To date, white loquat varieties are mainly derived from three pathways: firstly, the white meat is directly selected from white meat germplasm resources through actual seed selection, and most white meat varieties are derived from the way; secondly, breeding by taking white meat varieties as parents through a hybridization breeding way, wherein a few white meat varieties are derived from the way; thirdly, it is found from bud mutation that very few white meat varieties are derived from this approach. In the way, plants obtained by sowing in the first two main white loquat breeding modes generally need to be flowering and fruiting after more than 5 years of childhood, and the plant characteristics of seedlings are affected by grafting or repeatedly applying high-concentration hormone.
Disclosure of Invention
The invention aims to provide a breeding method of white loquat and a method for shortening the childhood period, which improve the screening efficiency of the hybrid parents and the hybrid offspring of the white loquat, the preservation time and pollination efficiency of loquat pollen, the yield of hybrid seeds, avoid the influence of grafting or repeated application of high-concentration hormone on the plant properties of the hybrid offspring of the loquat, and shorten the childhood period of the hybrid offspring of the loquat by more than 1 year.
The invention provides a breeding method of white loquat, which comprises the following steps: (1) With white loquat germplasm resource or EjPSY2A d The germplasm resource of the yellow flesh loquat of the gene is used as a parent;
(2) Pollinating the collected pollen of the male parent to the emasculated female parent, and bagging;
(3) The loquat seeds are collected in the next year sowing bags, and the seedlings identified as the white loquat germplasm are used for nursery establishment and field planting by a molecular marking method after emergence.
Preferably, in the step (1), the molecular marker primer is utilized to carry out PCR amplification on the germplasm resource of the eriobotrya japonica, and when the PCR amplification product shows a band with the length of 300-400 bp, the eriobotrya japonica is white; when the amplified product shows two bands of 300-400 bp and more than 500bp, the amplified product is the germplasm of the loquat with yellow flesh;
the molecular marker primer pair comprises PSY2A-2F and PSY2A-2R, wherein the nucleotide sequence of the PSY2A-2F is shown as SEQ ID NO.1, and the nucleotide sequence of the PSY2A-2R is shown as SEQ ID NO. 2.
Preferably, the PCR amplification procedure comprises: pre-denaturation at 94℃for 4min; denaturation at 95℃for 1min, annealing at 55℃for 45s, extension at 72℃for 1min, and cycling for 35 times; extending at 72℃for 10min.
Preferably, the collecting method of pollen in step (2) comprises: and (3) completely pinching off petals and stamens at the upper half part of the unopened flower bud, taking out, filtering to obtain stamens, and drying at constant temperature in a greenhouse at 25-28 ℃ for 24-36 h to obtain pollen.
Preferably, after pollen is obtained, the pollen is mixed with a neutral drying agent for preservation.
Preferably, the method of molecular marking in the step (3) comprises the steps of carrying out PCR amplification by using PSY2A-2F and PSY2A-2R, wherein when the PCR amplification product shows a band with the length of 300-400 bp, the product is white loquat germplasm;
the nucleotide sequence of PSY2A-2F is shown as SEQ ID NO.1, and the nucleotide sequence of PSY2A-2R is shown as SEQ ID NO. 2.
The invention also provides a method for shortening the childhood period of white loquat raised by the breeding method, which comprises the following steps: planting the seedlings of the white loquat on a high ridge of a nursery, and covering a film on the ridge surface after the planting; the height of the high ridge is 50cm, and the width of the ridge bottom is 2m;
topdressing nitrogen fertilizer during the tip pulling in the same year of field planting, and topdressing nitrogen, phosphorus and potassium fertilizer in the second year and the third year after field planting;
in the current year of field planting, 50-60 cm of stem is fixed, 3-4 main branches are selected and reserved, 2-3 side branches are selected and reserved on each main branch, and branch pulling is carried out in the second year after field planting;
the trunk is subjected to girdling in the third year after field planting, and the flower-promoting agent is sprayed in the fourth year; the flower promoting agent comprises monopotassium phosphate, borax and magnesium sulfate.
Preferably, the branch pulling comprises branch pulling in 7 late 7 th year after field planting, so that the angle between the main branch and the open branch of the central trunk is 60-80 degrees.
Preferably, the loquat is girdled for 3 weeks on the dry weight of the loquat in the third 6 months after field planting.
Preferably, the flower-promoting agent is sprayed 3 times from 6 months to 7 months of the fourth year after the planting.
The beneficial effects are that: the invention provides a breeding method of white-meat loquat and a method for shortening the childhood phase of the bred white-meat loquat, which specifically comprise the following steps: s1 parent selection, S2 pollen preparation, S3 hybridization assembly, S4 cold protection and fruit preservation, S5 sowing and seedling cultivation, S6 nursery establishment, S7 transplanting and field planting, S8 sapling fertilization, S9 sapling shaping, S10 comprehensive childhood shortening technology and the like. The method improves the screening efficiency of the hybrid parents and the hybrid offspring of the white-meat loquat, improves the preservation time and pollination efficiency of the loquat pollen, improves the yield of hybrid seeds, avoids the influence of grafting or repeated application of high-concentration hormone on the plant characters of the hybrid offspring of the loquat, shortens the childhood period of the hybrid offspring of the loquat by more than 1 year, and can lead the flowering rate of the hybrid offspring of the loquat after sowing to be more than 77.5 percent at the 4 th year.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions of the prior art, the drawings that are needed in the embodiments will be briefly described below, it being obvious that the drawings in the following description are only some embodiments of the present invention, and that other drawings may be obtained according to these drawings without inventive effort for a person skilled in the art.
FIG. 1 is a method for improving a civil engineering garden by adopting a soil gathering and ridging mode;
FIG. 2 is a method of pruning a post-field sapling;
fig. 3 shows the effect of promoting flowering by pulling branches, wherein the lower left corner of the picture indicates shooting time, place and longitude and latitude, which are 2021, 11 and 8, and are three sections of City, east mountain great road, 30.369826 degrees N and 104.157098 degrees E, and the same applies below;
fig. 4 is a comprehensive flower-promoting effect display, from top to bottom and from left to right, and each graph is labeled with the following information: 13-2 x high-ranked imperial concubine-54 (large flower quantity); huang Feng big five stars-6 (flowers big tree vigor middle); huang Feng big five stars-3 (big flower quantity); 43-15-12 (dwarfing and high flower size); white sand x Huang Mi 18 (large flower amount); white sand x large five stars-44 and 45 (large flowers); white sand x large five stars-25 (flower quantity is extremely large); large five stars x white sand-3 (flower quantity is extremely large); white sand X Huang Mi-17 (large flower amount); brilliant red x large five stars-4 (easy flower formation, large flower quantity); large five stars x white sand-7 (flowers rate almost 100%); large five stars x white sand-8 (flower formation rate almost 100%).
Detailed Description
The invention provides a breeding method of white loquat, which comprises the following steps: (1) With white loquat germplasm resource or EjPSY2A d The germplasm resource of the yellow flesh loquat of the gene is used as a parent;
(2) Pollinating the collected pollen of the male parent to the emasculated female parent, and bagging;
(3) The loquat seeds are collected in the next year sowing bags, and the seedlings identified as the white loquat germplasm are used for nursery establishment and field planting by a molecular marking method after emergence.
The invention selects excellent white loquat germplasm resources or selects EjPSY 2A-containing materials by using a loquat pulp color molecular marker identification technology according to breeding targets d The excellent yellow flesh loquat germplasm resource of the gene is used as a parent. In the invention, when the loquat pulp color molecular marker is used for identification, preferably, genomic DNA of loquat germplasm is used as a template, a PCR reaction system is prepared by using the genomic DNA and a molecular marker primer pair, PCR amplification is carried out, and genotyping is carried out on a PCR amplified product; when the PCR amplified product shows a band with the length of 300-400 bp, the loquat germplasm is white meat; when the PCR amplified product shows two bands of 300-400 bp and > 500bp, it contains EjPSY2A d The germplasm of the yellow flesh loquat of the gene (also can be used as a parent for hybridization combination). The reaction system for PCR amplification according to the present invention preferably comprises, in 25. Mu.L: 1.0. Mu.L template, 12.5. Mu.L 2 XPCR Mix, 1.0. Mu.L 10. Mu.M F primer, 1.0. Mu.L 10. Mu.M R primer and the balance ddH 2 O. The PCR amplification procedure of the present invention preferably comprises: pre-denaturation at 94℃for 4min; denaturation at 95℃for 1min, annealing at 55℃for 45s, extension at 72℃for 1min, and cycling for 35 times; extending at 72℃for 10min. After the amplification product is obtained in the present invention, the PCR amplification product is preferably genotyped using 8% polyacrylamide gel electrophoresis. The nucleotide sequence of PSY2A-2F is shown in SEQ ID NO. 1: AGCCTTTTCTTGAATGTGGTCG the nucleotide sequence of PSY2A-2R is shown in SEQ ID NO. 2: TTGATTGATGTTGGCTAATGAGATC. In the present invention, ejPSY2A is contained as identified by the method d The genetic yellow flesh loquat germplasm resource preferably comprises: oak leaf loquat, early red 89-4, huangfeng, xiangzhi No. 11, late clock 518, peyou, sichuan No.2, wulila, sichuan No. 4, early clock 6, mao mu, ji, taicheng No. 4, huang Jinkuai, jialing and Dahongpao.
Pollen is prepared when loquat buds are not bloomed, for example, in the embodiment of the invention, the un-bloomed loquat buds are collected for 1-2 times in autumn and winter in the adult region, petals and stamens at the upper half part of the flower are all pinched off and taken out by forceps, stamens are obtained after step-by-step filtration by 20-mesh and 35-mesh screens (the aperture of the screens can be adjusted according to the sizes of stamens of different loquat varieties), the stamens are placed in a greenhouse with the temperature of 25-28 ℃ for constant temperature drying of 24-36 h, the collected pollen is poured into a 50mL centrifuge tube, and a proper amount of neutral drying agent (anhydrous calcium chloride) or color-changing silica gel is added for cold storage at the temperature of 4 ℃. The pollen of the present invention is preferably derived from all of the above-mentioned parents, and is subjected to inter-parent cross.
The hybrid combination is carried out from the initial flowering stage to the full flowering stage of the female parent, and the pollination effect is good preferably from 10 to 17 times of windless clear weather. When the hybridization assembly is carried out, flowers which are already bloomed or not yet exposed are firstly removed, the artificial emasculation is carried out, the pollen before pollination is required to be placed under warm sunlight or activated by body temperature for 0.5-1 h, the whole inflorescence is immediately covered by a big fruit bag of 35 cm multiplied by 25 cm after pollination, and parent hybridization information is registered by hanging.
The invention preferably also comprises cold-proof fruit protection after artificial pollination, more preferably covers the tree tray with straw before low temperature freezing injury, improves the soil temperature, and simultaneously uses 7.2m wide cold-proof non-woven fabric (40 g/m in the planting row direction 2 ) Covering the whole tree crown.
The loquat seeds are harvested in the next year sowing bags according to the invention, the sowing substrate preferably comprises a mixture of peat, perlite and vermiculite, and the volume ratio of peat, perlite and vermiculite in the substrate is preferably 3:1:1. In the embodiment of the invention, the sowing is preferably performed in a seedling-raising square tray, the length of the seedling-raising square tray is preferably 42 x 5.5cm, 25 seeds are uniformly sown in each tray, and the greenhouse seedlings are grown in winter.
After the seeds sprout, screening of the hybrid offspring of the white-pulp loquat is carried out, preferably, PCR amplification is carried out by using PSY2A-2F and PSY2A-2R, and the judgment of the germplasm type is carried out according to the amplified products, wherein the procedure and the system of the PCR amplification are preferably the same as the above, and are not repeated here. The PCR amplification of the invention preferably comprises 45-60 d after sowing, taking young leaves or stem tips of seedlings for molecular marker identification, screening out white loquat hybrid offspring with 300-400 bp PCR amplification products, and listing for nursery establishment and concentrated field planting.
The invention preferably adopts a soil gathering and ridging mode to change a soil nursery, and comprises the steps of removing attachments on the soil in the garden before soil changing, leveling the soil, paying off according to planting row spacing of 4m multiplied by 5m, then intensively spreading base fertilizer in a ridging range, intensively piling surface soil between rows to a ploughed area after ploughing, and forming a tile-back ridge surface with the ridge height of about 50cm and the ridge bottom width of about 2 m. The base fertilizer of the invention preferably comprises decomposed organic fertilizer and superphosphate, wherein the dosage of the decomposed organic fertilizer is preferably 1000kg/667m 2 The dosage of the superphosphate is preferably 100kg/667m 2 Soil with the depth of 30 cm-40 cm is loosened by an excavator and evenly mixed with fertilizer, and then the surface soil between rows is intensively piled up in the ploughed area to form a tile-back-shaped ridge surface.
Before field planting, the invention preferably draws a field planting diagram according to different hybridization combinations, and transplants the nursery after the lowest air temperature in spring day is more than or equal to 10 ℃ (the field planting is suitable in the middle of 2 months to 3 months in Chengdu area). Before transplanting, 3-5 d spraying the leaves with broad-spectrum bactericide, and cutting off 1/2-1/3 of the leaf area of each leaf during seedling raising to reduce water evaporation. Digging a planting hole slightly larger than the root width of the hybrid seedling on the planting point, lightly placing the seedling into the planting hole, righting, taking cured surface fine soil as planting soil, and compacting after filling the soil to enable the root system to fully contact with the soil and enable the root neck to be exposed out of the ground. After the field planting is finished, a water ring with the diameter of more than or equal to 80cm is made, root fixing water is poured fully, the exposed root system is covered by fine soil, and a mulching film with the thickness of more than or equal to 0.03mm is selected for ridge surface covering.
The invention also provides a method for shortening the childhood period of white loquat raised by the breeding method, which comprises the following steps: planting the seedlings of the white loquat on a high ridge of a nursery, and covering a film on the ridge surface after the planting; the height of the high ridge is 50cm, and the width of the ridge bottom is 2m;
topdressing nitrogen fertilizer during the tip pulling in the same year of field planting, and topdressing nitrogen, phosphorus and potassium fertilizer in the second year and the third year after field planting;
in the current year of field planting, 50-60 cm of stem is fixed, 3-4 main branches are selected and reserved, 2-3 side branches are selected and reserved on each main branch, and branch pulling is carried out in the second year after field planting;
the trunk is subjected to girdling in the third year after field planting, and the flower-promoting agent is sprayed in the fourth year; the flower promoting agent comprises monopotassium phosphate, borax and magnesium sulfate.
The seedling setting of the white loquat of the invention is preferably the same as the above, and will not be described here.
After the field planting, the loquat hybrid seedlings follow the principle of applying Bao Shi on duty, preferably comprises the steps that the field planting can be applied once after the seedling is pulled for 15d in the spring, summer, autumn and winter each time, 100g of urea or high nitrogen water-soluble fertilizer is applied to each plant each time, the proportion of phosphorus and potassium fertilizers is gradually increased for the second and third years after the field planting, the flower bud differentiation is promoted, the fertilization time is consistent with the prior fertilization time, and the application is applied once after the seedling is pulled for 15d in the spring, summer, autumn and winter each time; if 50g of potassium dihydrogen phosphate is applied every time in the second year after field planting, the dosage reaches 100g in the third year.
After the young tree is planted, the invention also comprises young tree shaping, preferably comprising: the method of wiping off tender tips is mainly adopted in the current year after field planting of hybrid seedlings, redundant new tips are wiped off, and tips with proper directions and positions are selected and reserved. And (3) setting the dry length to be 50 cm-60 cm, selecting and reserving 3-4 main branches, and selecting and reserving 2-3 side branches for each main branch, wherein the side branches are uniformly distributed. In the invention, the branches are pulled in the next 7 th month after the field planting, so that the opening branch angle between the main branches and the central trunk is 60-80 degrees, and the side branches and the auxiliary branches are twisted to promote flower formation.
The invention carries out girdling on the dry weight of the loquat for 3 weeks in the third year 6 months after field planting, sprays '0.3 percent (m/m) monopotassium phosphate +0.3 percent (m/m) borax +0.1 percent (m/m) magnesium sulfate' 3 times in the fourth year 6 months to 7 months, sprays once every 10 days, combines the ridging drainage and ridge surface coverage in the earlier stage, reduces the soil humidity (average control is 15 to 18 percent), and can lead the flower forming rate of the 4 th year after the loquat filial generation sowing to be more than 77.5 percent.
For further explanation of the present invention, a method for breeding white loquat and a method for shortening childhood according to the present invention will be described in detail with reference to the accompanying drawings and examples, but they should not be construed as limiting the scope of the present invention.
Example 1
(1) Parent selection: selecting excellent white loquat germplasm resource or screening EjPSY 2A-containing loquat pulp color molecular marker identification technology (SEQ ID NO.1 and SEQ ID NO. 2) developed by utilizing project group according to breeding targets d The excellent yellow flesh loquat germplasm resource of the gene is used as a parent;
wherein EjPSY2A is contained d Genetic yellow flesh loquat germplasm resource: oak leaf loquat, early red 89-4, huangfeng, xiangzhi No. 11, late clock 518, peyou, sichuan No.2, wulila, sichuan No. 4, early clock 6, mao mu, ji, taicheng No. 4, huang Jinkuai, jialing and Dahongpao.
White loquat germplasm resource: soft white sand, sichuan No. 3, xinbai No. 8, xiangfei, huang Mi, guanyu, ninghai white, royal, baijiali No. 5, baijiali No. 7, xinbai No. 7, dabai pear, qingzhang, baiyu, chicken protein, taiping white, hua Bai No.1, feng Yu, jinyu, bai Maomu, wu Gongbai, rock candy seed, zaojia No. 5.
The genomic DNA is derived from tender leaves or stem tips of loquat germplasm;
the PCR amplification procedure includes: pre-denaturation at 94℃for 4min; denaturation at 95℃for 1min, annealing at 55℃for 45s, extension at 72℃for 1min, and cycling for 35 times; extending at 72℃for 10min.
The PCR reaction system is calculated by 25 mu L and comprises: 1.0. Mu.L template, 12.5. Mu.L 2 XPCRMIX, 1.0. Mu.L 10. Mu.M F primer, 1.0. Mu.L 10. Mu.M R primer and the balance ddH 2 O, genotyping the PCR amplification product using 8% polyacrylamide gel electrophoresis.
When the PCR amplified product shows a band with the length of 300-400 bp, the loquat germplasm is white meat; when the PCR amplified product shows two bands of 300-400 bp and > 500bp, it contains EjPSY2A d The germplasm of the yellow flesh loquat of the gene.
(2) Preparing pollen: collecting the un-bloomed loquat buds in autumn and winter in the Chengdu area for 1-2 times, taking out the petals and stamens at the upper half part of the flowers by pinching off with forceps, filtering step by step through a 20-mesh and 35-mesh screen, drying the stamens at constant temperature in a greenhouse at 25-28 ℃ for 24-36 hours, pouring the collected pollen into a 50mL centrifuge tube, adding a proper amount of neutral drying agent (anhydrous calcium chloride) or color-changing silica gel, and preserving at 4 ℃ in a refrigerated manner;
(3) Hybridization assembly: and the pollination effect is good when the windless clear weather from the initial flowering period to the full flowering period of the female parent is selected from 10 to 17. Firstly, flowers which are already bloomed or not yet exposed are removed, the flowers are artificially emasculated, pollen is required to be placed under warm sunlight or activated for 0.5 to 1 hour at body temperature before pollination, and the whole inflorescence is covered by a big fruit bag of 35 cm multiplied by 25 cm immediately after pollination;
(4) Cold-proof fruit-protection: before low temperature freezing injury, the tree plate is covered with straw to raise soil temperature, and 7.2m wide cold-proof non-woven fabric (40 g/m in the planting row direction 2 ) Covering the whole tree crown;
(5) Sowing and seedling raising: sowing the obtained seeds in seedling square trays with the length of 42 x and the width of 42 x and the height of 5.5cm in 7 months of the next year, wherein the seedling matrixes are peat, perlite and vermiculite=3:1:1, and 5*5 seeds are uniformly sown in each tray, so that greenhouse seedling can survive winter;
(6) Screening the white loquat hybrid offspring: and (3) taking young leaves or stem tips of seedlings for molecular marker identification 45-60 days after sowing by using the method in (1), screening out white loquat hybrid offspring with a PCR amplification product of 300-400 bp, and listing for nursery establishment and concentrated field planting. The hybrid 674 strain is obtained by accumulation, wherein 451 strain of white loquat plants are selected.
(7) And (3) nursery establishment: and (5) changing a soil-gathering ridging mode into a soil-building garden. Removing attachments on the land in the garden before soil improvement, leveling the land, paying off according to the planting line spacing of 4m multiplied by 5m, and then intensively broadcasting 1000kg/667m of decomposed organic fertilizer in the ridging range 2 +superphosphate 100kg/667m 2 Uniformly mixing soil with the depth of 30 cm-40 cm with fertilizer by using an excavator, and intensively stacking surface soil among rows to a ploughed area to form a tile-back ridge surface with the ridge height of about 50cm and the ridge bottom width of about 2m (figure 1);
(8) Transplanting and planting: firstly, initially drawing a field planting diagram according to different hybridization combinations, and transplanting a nursery after the lowest air temperature in spring day is more than or equal to 10 ℃ (the field planting is suitable in the middle of 2 months to 3 months in the Chengdu region). Before transplanting, 3-5 d spraying the leaves with broad-spectrum bactericide, and cutting off 1/2-1/3 of the leaf area of each leaf during seedling raising to reduce water evaporation. Digging a planting hole slightly larger than the root width of the hybrid seedling on the planting point, lightly placing the seedling into the planting hole, righting, taking cured surface fine soil as planting soil, and compacting after filling the soil to enable the root system to fully contact with the soil and enable the root neck to be exposed out of the ground. After the field planting is finished, making a water ring with the diameter of more than or equal to 80cm, watering enough root fixing water, covering the exposed root system with fine soil, and covering the ridge surface with a mulching film with the thickness of more than or equal to 0.03 mm;
(8) Fertilizing young trees: the seedling of the loquat hybrid seeds follows the principle of applying Bao Shi, the field planting can be carried out once about 15d after the seedling is pulled out each time in spring, summer, autumn and winter in the current year, 100g of urea or high-nitrogen water-soluble fertilizer is applied to each plant each time, the proportion of the phosphorus fertilizer and the potassium fertilizer is gradually increased in the second and third years after the field planting, and the flower bud differentiation is promoted;
(9) Comprehensive childhood shortening technology: the method of wiping off tender tips is mainly adopted in the current year after field planting of hybrid seedlings, redundant new tips are wiped off, and tips with proper directions and positions are selected and reserved. And (3) fixing the trunk to 50 cm-60 cm, and selecting 3-4 main branches. 2-3 side branches are reserved on each main branch, and are uniformly distributed. Pulling branches in 7 late days of the second year after field planting to enable the opening branch angle of the main branches and the central trunk to be 60-80 degrees, and twisting tips for the side branches and the auxiliary branches to promote flower formation (figures 2 and 3);
and after field planting, the main dry weight of the loquat is girdled for 3 weeks in 6 months in the third year, tree vigor is controlled, 3 times of '0.3% monopotassium phosphate +0.3% borax +0.1% magnesium sulfate' are sprayed from 6 months to 7 months in the fourth year, and the soil humidity is reduced by combining fertilization and shaping, ridging drainage and ridge surface coverage of young trees in the earlier stage, so that the flower forming rate of the loquat in the 4 th year after the hybrid offspring sowing is over 77.5 percent (figure 4).
The invention improves the screening efficiency of the hybrid parents of the white loquat, improves the preservation time and pollination efficiency of the loquat pollen, improves the yield of hybrid seeds, avoids the influence of grafting or hormone treatment on the plant characters of the hybrid offspring of the loquat, and shortens the juvenile period of the hybrid offspring of the loquat by more than 1 year.
Although the foregoing embodiments have been described in some, but not all, embodiments of the invention, it should be understood that other embodiments may be devised in accordance with the present embodiments without departing from the spirit and scope of the invention.
Claims (9)
1. The breeding method of the white loquat is characterized by comprising the following steps of: (1) With white loquat germplasm resource or EjPSY2A d The germplasm resource of the yellow flesh loquat of the gene is used as a parent;
(2) Pollinating the collected pollen of the male parent to the emasculated female parent, and bagging;
(3) The loquat seeds are collected in the next year sowing bags, and the seedlings identified as the white loquat germplasm are used for nursery establishment and field planting by a molecular marking method after emergence.
2. The breeding method according to claim 1, wherein in the step (1), the molecular marker primer is used for PCR amplification of the eriobotrya japonica germplasm resource, and when the PCR amplification product shows a band with the length of 300-400 bp, the eriobotrya japonica germplasm is white; when the amplified product shows two bands of 300-400 bp and more than 500bp, the amplified product is the germplasm of the loquat with yellow flesh;
the molecular marker primer pair comprises PSY2A-2F and PSY2A-2R, wherein the nucleotide sequence of the PSY2A-2F is shown as SEQ ID NO.1, and the nucleotide sequence of the PSY2A-2R is shown as SEQ ID NO. 2.
3. The method of breeding according to claim 1, wherein the method of collecting pollen in step (2) comprises: and (3) completely pinching off petals and stamens at the upper half part of the unopened flower bud, taking out, filtering to obtain stamens, and drying at constant temperature in a greenhouse at 25-28 ℃ for 24-36 h to obtain pollen.
4. A method of breeding according to claim 3, further comprising, after obtaining pollen, mixing the pollen with a neutral desiccant for preservation.
5. The breeding method according to claim 1, wherein the method of molecular marker in the step (3) comprises performing PCR amplification using PSY2A-2F and PSY2A-2R, and the white loquat germplasm when the PCR amplification product shows a band of 300-400 bp in length;
the nucleotide sequence of PSY2A-2F is shown as SEQ ID NO.1, and the nucleotide sequence of PSY2A-2R is shown as SEQ ID NO. 2.
6. A method for shortening the childhood period of white-fleshed loquat raised by the breeding method of claims 1 to 5, comprising the steps of: planting the seedlings of the white loquat on a high ridge of a nursery, and covering a film on the ridge surface after the planting; the height of the high ridge is 50cm, and the width of the ridge bottom is 2m;
topdressing nitrogen fertilizer during the tip pulling in the same year of field planting, and topdressing nitrogen, phosphorus and potassium fertilizer in the second year and the third year after field planting;
in the current year of field planting, 50-60 cm of stem is fixed, 3-4 main branches are selected and reserved, 2-3 side branches are selected and reserved on each main branch, and branch pulling is carried out in the second year after field planting;
the trunk is subjected to girdling in the third year after field planting, and the flower-promoting agent is sprayed in the fourth year; the flower promoting agent comprises monopotassium phosphate, borax and magnesium sulfate.
7. The method of claim 6, wherein the pulling comprises pulling the branch in the second 7 th and last year after the planting such that the main branch opens at a branching angle of 60 ° to 80 ° from the central trunk.
8. The method of claim 6, wherein the loquat is girdled for 3 weeks on the dry weight of the loquat in the third 6 th year after the permanent planting.
9. The method of claim 6, wherein the flower-promoting agent is sprayed 3 times from 6 months to 7 months of the fourth year after the planting.
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