Efficient breeding method of new larch high-resistance gypsy moth germplasm

Abstract

The present application relates to a kind of efficient breeding methods of high resistance larch new germplasm of pine moth, including larch high resistance scion collection, rootstock selection and processing, grafting breeding and grafting management etc., in the process of high resistance scion seedling breeding, by scientific seed winter storage, sowing seedling and seedling management etc., to obtain a large number of healthy high resistance scion tender shoot provides guarantee. Scion uses the tender shoot of 10-15 days growth on seedling, and the high adaptability larch seedling of 10-15 days seedling stem straight thick, top bud full, root system developed, no disease and injury is selected as rootstock, so that grafting joint can heal quickly, shorten breeding cycle. After grafting, first grow in the environment very stable plant growth room for about 2 months, etc. Scion survival, transition period is transferred into greenhouse and grows for 1 year to reach healthy seedling, finally, further seedling raising in outdoor open-air test ground, so that seedling is fully lignified, thus a large number of high resistance pine moth larch seedling resources for artificial afforestation are obtained. The present application reduces the probability of occurrence of insect pests from the source, improves the insect resistance of larch itself, and improves the forest quality, growth rate and wood utilization efficiency.

Description

Technical Field

[0001] This invention relates to the field of forest resource maintenance technology, specifically to an efficient breeding method for a new germplasm of larch highly resistant to pine caterpillars. Background Technology

[0002] larch ( Larixgmelinii Larch (Ruprecht) Kuzeneva is a deciduous tree belonging to the genus Larch in the family Pinaceae. It can grow in a variety of environments, often forming large-scale pure stands or mixed forests dominated by larch, along with other coniferous and broad-leaved trees such as birch, black birch, bush birch, aspen, Scots pine, red spruce, and spruce. Larch thrives best in moist, well-drained, well-aerated, deep, and fertile soil. Larch wood possesses excellent mechanical properties, strong corrosion resistance, rapid growth, and strong tolerance to soil conditions and weather, making it a valuable material for transportation and construction. It can be used for railway sleepers, bridges, vehicle materials, mine props, and utility poles. Furthermore, turpentine, alcohol, and cellulose can be extracted from the wood, and the bark is used in leather making and dyeing. In summer, larch leaves exude a white, sweet substance called larch mannan, which contains a rare melitriose and has been used in clinical medicine.

[0003] However, the cultivation and growth of larch also face many problems, especially insect pests. Insect pests cause significant timber loss and reduced timber production in larch. The larch caterpillar is a very serious forest defoliant. For many years, traditional control measures have been used to reduce the damage caused by the larch caterpillar, including manual removal of cocoons and capture of larvae, and large-scale aerial spraying of pesticides. On the one hand, manual pest control is too labor-intensive and unsuitable for large-scale forest applications, while aerial spraying is costly, requiring regular, targeted spraying of pesticides in pine forests annually. On the other hand, pesticides cause significant environmental pollution and can easily induce highly resistant caterpillar species, thus rendering traditional control measures ineffective. Due to the increasingly serious larch caterpillar infestation, the breeding of new larch caterpillar-resistant germplasm is urgently needed. However, there are currently no reports on standardized and regulated propagation techniques for new larch germplasm highly resistant to larch caterpillars.

[0004] In view of this, it is necessary to formulate a set of efficient breeding strategies for larch with high resistance to pine caterpillars, so as to promote the industrialization of improved varieties and production engineering technology, accelerate the process of improved varieties, targeted production and industrialization of larch plantations, and improve the productivity of larch plantations. Summary of the Invention

[0005] (a) Technical problems to be solved

[0006] In view of the above-mentioned shortcomings and deficiencies of the existing technology, the present invention provides an efficient breeding method for a new larch germplasm highly resistant to pine caterpillars. The method uses asexual reproduction breeding to obtain a new larch germplasm highly resistant to pine caterpillars, thereby controlling larch pests at the source, accelerating the process of larch insect-resistant breeding, and applying the obtained new larch germplasm highly resistant to pine caterpillars to the large-scale construction of larch plantations. This provides a strong guarantee for the sufficient and sustainable supply of larch timber resources and solves the problems of high labor intensity, high cost, poor effectiveness, and environmental pollution of existing larch caterpillar control methods.

[0007] (II) Technical Solution

[0008] A highly efficient breeding method for a new larch germplasm highly resistant to pine caterpillars includes the following steps:

[0009] S1. Collection and storage of highly resistant timber

[0010] Excellent single trees with a morphological quality coefficient ≥80%, free from diseases, vigorous growth, and high insect resistance, aged 9-11 years, are used as seed-collecting mother trees. Cones are collected from the mother trees, treated for seed production, and seedlings are propagated in the nursery. Branches that have grown for 10-15 days are collected from the seedlings, retaining a 5-6 mm length of the branch below the leaves, and are cut at an oblique angle (preferably 45°) to obtain scions. The scions are stored in an insulated box at 20-25℃ and 70-80% humidity. The base of the scion is inserted 2-3 mm into a substrate treated with 40-60 mM IAA auxin for 50-70 minutes. The IAA auxin treatment substrate is prepared by dissolving IAA in 75% ethanol.

[0011] S2, Rootstock Treatment

[0012] Select 10-15 day old, straight and robust larch seedlings with plump terminal buds, well-developed root systems, and free from diseases, pests, and mechanical damage as rootstock. Cut a smooth surface at an angle (preferably 45°) from top to bottom on a smooth part of the hypocotyl of the rootstock needles, 0.5-0.7 cm from the base. Select rootstock of appropriate thickness according to the thickness of the scion, so that the thicknesses of the two are similar. The cut surface of the rootstock should be the same width as the cut surface of the scion, and the length of the cut surface of the rootstock should be 0.3-0.8 cm longer than the length of the cut surface of the scion.

[0013] S3, grafting and propagation

[0014] The slanted cut surface of the scion is joined with the slanted cut surface of the rootstock. After joining, the scion and rootstock are close together and the pith cambium is in close contact. Use waterproof tape to tie the scion and rootstock together, and the waterproof tape can completely cover and seal the cut surfaces of the scion and rootstock.

[0015] S4. Post-grafting management

[0016] Grafted larch seedlings were promptly transferred to a plant growth chamber for cultivation. The plant growth chamber operated on a 16h / 8h light-dark cycle, with a constant light intensity of 100-140 µmol / m². 2 • Keep the temperature at 20-25℃; after the scion sprouts new shoots, immediately remove the binding tape and water thoroughly once.

[0017] After one year of growth in a greenhouse, the seedlings were transplanted to outdoor experimental seedbeds. During the growth period in both the greenhouse and outdoor experimental seedbeds, watering was carried out promptly according to soil moisture conditions, and manual weeding was performed as needed. Outdoor cultivation was carried out from late June to mid-July of the same year, at a rate of 60-100 kg / hm². 2 Apply the urea solution evenly to the seedbed once.

[0018] According to a preferred embodiment of the present invention, in S1, the seed-collecting mother trees are from seed orchards or scion nurseries of various levels of superior seed sources, families, and clones suitable for the local area (proving that the scions have fast growth and high insect resistance at the growth and genetic levels); the cones are collected from the middle and rear parts of the current year's lignified branches with plump, healthy, and robust terminal buds on the outer side of the upper part of the larch crown.

[0019] The plump, healthy, and robust lignified branches from the upper outer part of the larch mother tree's crown possess ample nutrients and strong growth and healing abilities, effectively ensuring the full maturation and nutrient supply of the seeds within the collected cones. Cones from the middle and rear parts of the branches are collected for seed production, as the seeds in these cones are relatively mature and have a higher nutrient content.

[0020] The collected scions were stored in an insulated box with a mesh screen fixed in the lower middle part of the box. The scions were inserted into the mesh to secure them and prevent tilting. The base of the scion was immersed in a treatment substrate with a concentration of 50 mM IAA auxin for 1 hour to a depth of 2 mm. This ensured that the base of the seedlings did not lose moisture and that the hormone treatment promoted cell elongation and relaxation of the cell wall structure, thus improving the speed and quality of healing. The solvent for the treatment substrate was ethanol; therefore, the scions should not be immersed in the substrate for too long, otherwise it may cause changes in the cell wall structure.

[0021] In addition, plump and healthy semi-lignified branches from the current year can be collected from the mother tree as scions and grafted onto 1-3 year old highly adaptable larch seedling rootstocks to obtain larger larch seedlings highly resistant to pine caterpillars.

[0022] According to a preferred embodiment of the present invention, the lower end of the scion is cut at an angle of 45° to an elliptical surface. The smooth part of the rootstock at 0.5cm from the hypocotyl of the needle is cut into a smooth plane at an angle of 40-45° from top to bottom. After the scion's oblique cut is joined with the rootstock's cut, the scion and rootstock are brought close together to form a vertical stem. A silicone ring with a diameter of 0.1cm and a height of 0.6cm is used to bind the joint surface. When binding, the cut should be completely sealed to achieve the purpose of moisture retention.

[0023] According to a preferred embodiment of the present invention, in S4, the plant growth chamber is equipped with a 16h / 8h light-dark alternation, with the light intensity kept constant at 120µmol / m². 2 At a constant temperature of 23℃, the scions sprout new shoots after two months of growth in the plant growth chamber, indicating successful grafting. The stable environment of the plant growth chamber greatly improves the survival rate of the scions. After successful grafting, the binding tape is removed immediately, and the scions are transferred to a greenhouse for one year of growth before being transplanted to an outdoor experimental seedbed. Growing in the outdoor experimental seedbed allows the seedlings sufficient time to harden, ensuring adequate lignification and improving seedling survival rate. When weeding manually, hold the weeds down by pressing down on both sides of the roots to avoid uprooting the larch seedlings from the soil.

[0024] According to a preferred embodiment of the present invention, in order to improve the germination rate and quality of seedlings grown in the nursery and to increase the survival rate of grafted propagation, the method for cone seed production and nursery propagation of seedlings in step S1 is as follows:

[0025] Step 1: Cone Seed Production

[0026] Sun-drying the cones in the open air causes the seeds to fall off. Select the seeds carefully and dry them to a safe moisture content of 7-9%. Disinfect them by soaking them in a 0.3-0.5% potassium permanganate solution for 1-2 hours. Remove them, seal them for 20-40 minutes, rinse them with clean water, and then air-dry them for later use.

[0027] Step 2: Seed winter storage

[0028] Disinfected seeds are mixed with pure, moist sand at a mass ratio of 1:2.8-3.2 to obtain mixed sand seeds. The moisture content of the moist sand is 55-65% of saturated moisture (it can be formed into a ball by hand without releasing water, and it will not fall apart when released). First, a 10-15cm layer of moist sand is placed at the bottom of the pit, then the mixed sand seeds are spread out, and finally the surface is covered with a 15-25cm layer of fine moist sand. 3-5 bundles of straw or wheat straw with a diameter of 3.5-6.5cm are erected per square meter in the pit as ventilation holes. The straw or wheat straw bundles should be placed below the ground level at the bottom of the pit. The pit surface is covered with wire mesh to prevent rodent damage. After the seeds in the pit have frozen solid, a 25-45cm thick straw mat is placed over it for shade.

[0029] Step 3: Seedbed preparation

[0030] During land preparation, apply 5000-10000 kg of well-rotted organic fertilizer and 20-25 kg of superphosphate per mu as base fertilizer. 5-7 days before sowing, spray the seedbed surface with 0.4-0.5% potassium permanganate for soil disinfection. Mix 2-3 g of phoxim with an appropriate amount of fine soil per square meter for soil pest control. When constructing the seedbed, ensure that the seedbed surface is 10-20 cm higher than the walkway and leave drainage ditches between the seedbed surfaces. After the seedbed is formed, harrow and level it, and water it thoroughly before sowing.

[0031] Step 4: Sowing and Seedling Raising

[0032] Sowing time is after the soil thaws in spring, when the average daily temperature is stable at 11-14℃ and the temperature at a vertical depth of 10cm is stable at 7-10℃. 5-7 days before sowing, choose a sunny day, spread the cleaned winter-stored seeds evenly on the plastic sheet or canvas inside a temporary plastic greenhouse, with a seed layer thickness of 5-10cm. Keep the seed moisture at 55-65%. At night or on cloudy days, pile the seeds up and spread them out during the day, turning them over in time to prevent rotting. When at least 30% of the seeds show white sprouts and ≥50% of the seeds have turned their tips, it is ready for sowing.

[0033] The sowing method is as follows: sow in furrows with a sowing width of 9-12cm and a furrow depth of 0.4-0.7cm, cover with soil with a thickness of 0.3-0.5cm, compact with a manual roller after covering with soil, spray water to replenish the moisture of the seed layer, and add more soil to cover the seeds that are exposed after spraying water.

[0034] Step 5: Seedling Management

[0035] Seedlings emerge 15-20 days after sowing. During the emergence period, water 3-5 times a day, with each watering amount being 40-50 mL / m². Watering should be done before 9:00 AM and after 4:00 PM. Increase the watering frequency by 1-2 times during windy or hot weather. After emergence, spray 2-3 times a day, increasing the water amount to 80-100 mL / m² each time to encourage root growth.

[0036] (III) Beneficial Effects

[0037] The present invention includes the following steps: collection and storage of highly resistant larch scions, selection and treatment of rootstocks, grafting propagation, and post-grafting management. In the collection of highly resistant larch scions, seeds with relatively mature textures and high nutrient content are obtained by selecting mother trees and collection sites, providing a foundation for high germination rate and healthy seedlings. In the propagation of highly resistant scion seedlings, effective methods such as winter storage of seeds, sowing and seedling raising, and seedling management provide technical support for obtaining a large number of robust and usable highly resistant scion shoots, ensuring a high number of viable grafts and subsequent survival. In this invention, highly adaptable larch seedlings with straight, robust stems, full terminal buds, well-developed root systems, and free from pests, diseases, and mechanical damage (10-15 days old) are selected as rootstocks. This facilitates binding of the scion and rootstock and ensures close contact of the pith cambium, improving the scion survival rate and shortening the entire propagation and production cycle. After grafting, the seedlings are first grown in a very stable indoor plant growth chamber for about two months to allow the scions to sprout new shoots, which greatly improves the survival rate of the scions. During the transition period, they are moved to a greenhouse for one year to grow into robust seedlings. Finally, they are grown in an outdoor experimental seedbed to further harden the seedlings, ensuring that they are fully lignified and improving the seedling survival rate and survival rate. This allows for the efficient acquisition of a large number of highly resistant larch seedlings that can be used for artificial afforestation.

[0038] The present invention provides a method for breeding highly resistant larch trees to caterpillars. This method utilizes the seeds of a very limited number of disease-free, robust, and highly insect-resistant superior individual trees as breeding resources. By combining asexual reproduction breeding methods, a standardized and efficient breeding method for highly resistant new larch germplasm is obtained, which accelerates the insect-resistant breeding process of larch, controls pests at the source, and can be applied on a large scale to the establishment of larch plantations. This provides a strong guarantee for the sufficient and sustainable supply of larch timber resources and solves the problems of high labor intensity, high cost, poor effectiveness, and environmental pollution associated with existing larch caterpillar control methods. Detailed Implementation

[0039] To better explain and facilitate understanding of the present invention, the present invention will be described in detail below through specific embodiments.

[0040] Example 1

[0041] This embodiment describes an efficient breeding method for a new larch germplasm highly resistant to pine caterpillars, and the steps are as follows:

[0042] (I) Collection and storage of highly resistant scion wood

[0043] 1. Collect cones

[0044] ① Selecting mother trees for seed collection

[0045] Excellent larch trees of about 10 years old with fast growth, robust growth, high morphological coefficient (≥80%), no disease, and high insect resistance, suitable for the Lesser Khingan Mountains region, are selected from superior larch seed sources, families, clones, and seed orchards and scion nurseries at all levels. These trees are used as seed mother trees to ensure that the scions have fast growth and high insect resistance in terms of growth traits and genetic level.

[0046] ② Collect cones from the mother tree

[0047] Collect cones from the upper, middle, and lower parts of the current year's lignified branches on the outer, plump terminal buds of highly resistant caterpillar-bearing larch mother trees. These branches possess ample nutrients and strong growth and healing abilities, ensuring full seed maturation and nutrient supply. The cones from the upper, middle, and lower parts of these branches are reserved for seed production, as the seeds in these cones are relatively mature and have a higher nutrient content.

[0048] 2. Cone seed production and nursery propagation of seedlings

[0049] ① Cone seed production

[0050] Sun-drying the cones in the open air causes the seeds to fall off. Select the seeds carefully and dry them to a safe moisture content of 9%. Disinfect them by soaking them in a 0.5% potassium permanganate solution for 2 hours. Remove them, seal them for 30 minutes, rinse them with clean water, and then air-dry them for later use.

[0051] ② Seeds stored for winter

[0052] Sterilized seeds are mixed with high-quality, pure, moist sand at a mass ratio of 1:3 to obtain mixed sand seeds. The moisture content is 60% of saturation (the seeds should clump together when squeezed but not release water, and crumble easily when released). First, a 15cm layer of moist sand is placed at the bottom of the pit, then the mixed sand seeds are spread evenly, and finally, a 20cm layer of fine moist sand is placed on top. Four 4.5cm diameter bundles of wheat straw are erected per square meter in the pit as ventilation holes, with the straw bundles extending from the bottom of the pit to above ground level. The pit surface is covered with wire mesh to prevent rodent damage. After the seeds in the pit have frozen solid, a 30cm thick straw mat is placed over them for shade.

[0053] ③ Seedbed preparation

[0054] Combine land preparation with the application of a mixture of well-rotted organic fertilizer and chemical fertilizer as base fertilizer. Apply 7000 kg of well-rotted organic fertilizer and 20 kg of superphosphate per acre as base fertilizer. Five days before sowing, spray the seedbed surface with 0.5% potassium permanganate for soil disinfection. Mix 2 g of phoxim with an appropriate amount of fine soil per square meter for soil pest control. When constructing the seedbed, ensure that the seedbed surface is 15 m higher than the walkway, the seedbed surface is 120 cm wide, the walkway is 40 cm wide, and drainage ditches are left between the seedbed surfaces. After the seedbed is formed, finely rake and level it. Water thoroughly before sowing.

[0055] ④ Sowing and seedling raising

[0056] Sowing time is after the soil thaws in spring, when the average daily temperature is stable at 12℃ and the soil temperature at a vertical depth of 10cm is stable at 8℃. Five days before sowing, on a sunny day, spread the cleaned, winter-stored seeds evenly on a plastic sheet inside a temporary plastic greenhouse, with a seed layer thickness of 8cm. Maintain seed moisture at 60%. At night or on cloudy days, pile the seeds up and spread them out during the day, turning them regularly to prevent rotting. When at least 30% of the seeds show white sprouts and ≥50% of the seeds have turned from their seed coat to their buds, they are ready for sowing. Sowing method: Sow in rows with a sowing width of 10cm and a furrow depth of 0.5cm, evenly scattering the seeds in the furrows, and covering with 0.3-0.5cm of soil. After covering with soil, compact with a manual roller, spray water to replenish the moisture of the seed layer to catch up with the soil moisture, and cover any seeds exposed after watering with more soil.

[0057] ⑤ Seedling Management

[0058] Seedlings emerge 15-20 days after sowing. During the emergence period, water five times a day, with each application of 40 mL / m², using small, frequent applications. Watering should be done before 9:00 AM and after 4:00 PM. Increase the frequency by two times during windy or hot weather. After emergence, spray three times a day, increasing the water volume to 80 mL / m² each time to encourage root growth. After rain, promptly spray water to rinse the seedling stems to prevent sunburn. If the soil surface temperature exceeds 30℃, spray water to cool it down.

[0059] 3. Scion collection and pretreatment

[0060] ①Collecting scions

[0061] Collect tender branches from seedlings that are 15 days old to promote rapid healing at the grafting point. When collecting branches, retain 5mm of the branch below the leaves and make a 45° oval cut. Attach a label indicating the collection time and location.

[0062] ② Scion pretreatment

[0063] The collected young scions were stored in an insulated box at 25℃ and 80% humidity to prevent moisture loss. A mesh screen was placed in the lower middle part of the insulated box. The base of the scion was inserted through the mesh into a treatment substrate containing 50 mM indoleacetic acid (IAA). The mesh screen was fixed in the lower middle part of the insulated box to secure the scion and prevent tilting. The treatment substrate was prepared by dissolving IAA in 75% ethanol. The scion was submerged 2 mm in the treatment substrate for 1 hour to ensure that the base of the seedling did not lose moisture and that the hormone treatment promoted cell elongation and relaxation of cell wall structure, thus improving the speed and quality of healing.

[0064] (ii) Rootstock treatment

[0065] The choice of rootstock also affects the survival rate. In this example, 15-day-old, straight and robust larch seedlings with plump terminal buds, well-developed root systems, and free from pests, diseases, and mechanical damage were selected as rootstocks (highly adaptable larch seedlings refer to larch varieties suitable for planting in the target forest area, characterized by well-developed root systems, strong resistance, and outstanding target traits). A smooth plane was cut at a 45° angle from top to bottom on a smooth part 0.5 cm from the hypocotyl of the needles on the rootstock. When selecting rootstocks, rootstocks of appropriate thickness were chosen based on the thickness of the scion, ensuring that the thicknesses of the two are similar. The width of the cut surface of the rootstock is the same as that of the cut surface of the scion, and the length of the cut surface of the rootstock is 0.1 m longer than that of the cut surface of the scion.

[0066] In the Lesser Khingan Mountains region, the most adaptable larch species is the Dahurian larch (Larix gmelinii), therefore, Dahurian larch seedlings were selected as rootstock in this embodiment. Dahurian larch has the following characteristics: ① Rapid growth and adaptation to a short growing season: Dahurian larch grows rapidly, making full use of the short growing season. ② Wide soil adaptability: It can grow in a variety of soil types, from deep fertile to thin breccia layers and swamps, demonstrating strong environmental adaptability. ③ Cold hardiness: As a cold-temperate tree species, Dahurian larch is particularly cold-hardy and adapted to the cold climate conditions of the Lesser Khingan Mountains.

[0067] (III) Grafting and Propagation

[0068] Align the scion's oblique cut with the rootstock's oblique cut, ensuring the scion and rootstock are close together and their pith cambium layers are in close contact. Securely bind them together with a silicone ring with a diameter of 0.1cm and a height of 0.6cm, ensuring the cut is completely sealed during binding to maintain moisture.

[0069] (iv) Post-grafting management

[0070] Grafted larch seedlings were promptly transferred to a plant growth chamber for cultivation. The plant growth chamber operated on a 16h / 8h light-dark cycle, with a constant light intensity of 120 µmol / m². 2 •s, maintain the temperature at 23℃. After about 2 months in the plant growth room, when the graft union is completely healed and new shoots emerge from the scion (indicating successful grafting), immediately remove the binding material and water thoroughly once. Then transfer to a greenhouse (polytunnel) for 1 year of growth, and then transplant to an outdoor experimental seedbed.

[0071] During the growth period in greenhouses and outdoor open-field experimental seedbeds, watering should be done promptly according to soil moisture. Weeding should be done manually as needed, pressing the weeds down by the sides to avoid uprooting the larch seedlings. In outdoor open-field experimental seedbeds, from late June to mid-July of the same year, apply a urea solution evenly to the seedbed at a rate of 75 kg / hm². Transplanting to the outdoor experimental field allows the seedlings sufficient time to harden, ensuring adequate lignification and improving seedling survival rate.

[0072] Example 2

[0073] The difference between this embodiment and Embodiment 1 is that the seed-collecting mother trees are 11-year-old, fast-growing, robust, high-morphological-quality-index (≥80%), disease-free, and highly insect-resistant superior individual plants from the scion orchard. The remaining steps are the same as in Embodiment 1.

[0074] Example 3

[0075] The difference between this embodiment and Embodiment 1 is that the base of the young scion was inserted through a mesh into a treatment substrate with an IAA auxin concentration of 55 mM, to a depth of 2 mm, for a treatment time of 55 min. During post-grafting management, the plant growth chamber was kept under alternating light and dark conditions for 16 h / 8 h, with a constant light intensity of 110 µmol / m². 2 The plants were kept at a temperature of 25°C and grown in the plant growth chamber for two months before being moved to the greenhouse. The remaining steps were the same as in Example 1.

[0076] Comparative Example 1

[0077] The difference between this comparative example and Example 1 is that, in the process of propagating seedlings in the nursery, the seeds were not spread out and turned over in advance to awaken them. Instead, the seeds were directly taken out and sown in furrows. The sowing method was the same as in Example 1.

[0078] Comparative Example 2

[0079] The difference between this comparative example and Example 1 is that the scions collected were branches from seedlings that had grown for 40 days. The scion cutting and pretreatment, as well as other treatment steps and methods, were the same as in Example 1.

[0080] Comparative Example 3

[0081] The difference between this comparative example and Example 1 is that the grafted seedlings were not allowed to grow in a greenhouse for one year after grafting; instead, they were transplanted to an outdoor experimental seedbed shortly after the scion emerged. All other treatment steps and methods were the same as in Example 1.

[0082] Cones were collected from the upper and lower outer terminal buds of the current year's lignified branches of highly resistant larch mother trees, taken from the middle and rear parts of the outermost part of the crown. These cones were dried to obtain 1000 plump, undamaged larch seeds. Seedlings of highly resistant larch were then cultivated using the methods described in Examples 1-3 and Comparative Examples 1-3. The germination rate of seedlings from the highly resistant larch nursery and the number of seedlings ultimately usable for plantation establishment in the outdoor experimental seedbeds were statistically analyzed. The results are as follows:

[0083]

[0084] The results above show that, under standardized procedures for seed production from highly resistant larch cones and seedling propagation in nurseries, the number of seedlings produced in nurseries remains relatively stable, except for a lower germination rate in Comparative Example 1. However, when grafting to propagate highly resistant larch seedlings suitable for plantation, many factors can change, including the selection of scions and rootstocks, and post-grafting management. Failure to follow the standardized procedures described in this invention will directly lead to a significant decrease in the seedling survival rate of highly resistant larch seedlings, affecting seedling production efficiency and increasing the cost of propagating highly insect-resistant seedlings.

[0085] The breeding method of this invention can obtain a new variety of larch with high resistance to pine caterpillars in a short period of time, reducing the probability of pest occurrence from the source, improving the larch's own resistance to pests, and improving forest quality and timber utilization efficiency. The breeding method has a short production cycle, low production cost, small land area, and high survival rate, and has good practical application prospects. It can be applied to the construction of larch timber forests in northern my country to reduce the loss of larch timber caused by pine caterpillar pests.

[0086] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A highly efficient breeding method for a new larch germplasm highly resistant to pine caterpillars, characterized in that, Includes the following steps: S1. Collection and storage of highly resistant timber Excellent single trees with a morphological quality coefficient ≥80%, free from disease, vigorous growth, and high insect resistance, aged 9-11 years, are used as seed-collecting mother trees. Cones are collected from the mother trees, treated for seed production, and seedlings are propagated in the nursery. Branches that have grown for 10-15 days are collected from the seedlings, retaining a 5-6 mm length below the leaves, and scions are obtained by oblique cuts. The scions are stored in an insulated box at 20-25℃ and 70-80% humidity. The base of the scions is inserted 2-3 mm into a substrate treated with 40-60 mM IAA auxin for 50-70 minutes. The IAA auxin treatment substrate is prepared by dissolving IAA in 75% ethanol. The cones are collected from the middle and rear parts of the plump, healthy, and robust current-year lignified branches on the outer side of the upper-middle part of the larch crown; the seed production method from the cones is as follows: Sun-drying the cones in the open air causes the seeds to fall off. Select the seeds carefully and dry them to a safe moisture content of 7-9%. Disinfect them by soaking them in a 0.3-0.5% potassium permanganate solution for 1-2 hours. Remove them, seal them for 20-40 minutes, rinse with clean water, and then air-dry them until ready for use. Mix the disinfected seeds with pure, moist sand at a mass ratio of 1:2.8-3.2 to obtain a mixed sand seed mixture. The moisture content of the moist sand should be 55-65% of its saturated moisture content. First, lay a 10-15cm layer of moist sand at the bottom of a pit, then spread the mixed sand seed mixture evenly. After the seeds are frozen solid, cover the surface with 15-25cm of fine, damp sand. Place 3-5 straw or wheat straw bundles with a diameter of 3.5-6.5cm per square meter in the pit as ventilation holes, extending the straw or wheat straw bundles down to the bottom of the pit above the ground. Cover the pit surface with wire mesh to prevent rodent damage. After the seeds have solidified in the pit, cover them with a 25-45cm thick straw mat for shade. Once the soil thaws in spring, remove the seeds and sow them in a pre-prepared seedbed with well-rotted organic fertilizer and superphosphate, and after soil disinfection and pest control treatment, to prepare for seedling propagation. S2, Rootstock Treatment Select 10-15 day old, straight and robust larch seedlings with plump terminal buds, well-developed root systems, and free from pests, diseases, and mechanical damage as rootstock. Cut a smooth surface at an angle from top to bottom on a smooth part of the hypocotyl of the rootstock needles, 0.5-0.7 cm from the base. Select rootstock of appropriate thickness according to the thickness of the scion, so that the thickness of the two is similar. The cut surface of the rootstock should be the same width as the cut surface of the scion, and the length of the cut surface of the rootstock should be 1-3 mm longer than the length of the cut surface of the scion. S3, grafting and propagation The slanted cut surface of the scion is joined with the slanted cut surface of the rootstock. After joining, the scion and rootstock are close together and the pith cambium is in close contact. Use waterproof tape to tie the scion and rootstock together, and the waterproof tape can completely cover and seal the cut surfaces of the scion and rootstock. S4. Post-grafting management Grafted larch seedlings were promptly transferred to a plant growth chamber for cultivation. The plant growth chamber operated on a 16h / 8h light-dark cycle, with a constant light intensity of 100-140 µmol / m². 2 • Keep the temperature at 20-25℃; after the scion sprouts new shoots, immediately remove the binding tape and water thoroughly once. After one year of growth in a greenhouse, the seedlings were transplanted to outdoor experimental seedbeds. During the growth period in both the greenhouse and outdoor experimental seedbeds, watering was carried out promptly according to soil moisture conditions, and manual weeding was performed as needed. Outdoor cultivation was carried out from late June to mid-July of the same year, at a rate of 60-100 kg / hm². 2 Apply the urea solution evenly to the seedbed once.

2. The efficient breeding method according to claim 1, characterized in that, In S1, the seed-collecting mother trees come from seed orchards or scion nurseries of various levels that are suitable for the region, and are of superior seed sources, families, or clones.

3. The efficient breeding method according to claim 1, characterized in that, In S1, a mesh is fixed in the lower middle part of the incubator. The scion is inserted into the mesh to fix it and prevent it from tilting. The base of the scion is immersed in a treatment substrate with an IAA auxin concentration of 50mM for 1 hour.

4. The efficient breeding method according to claim 1, characterized in that, The lower end of the scion is cut at a 45° elliptical angle. The smooth part of the rootstock, 0.5cm from the hypocotyl of the needle, is cut at a 40-45° angle from top to bottom to form a smooth plane. After the scion's cut is aligned with the rootstock's cut, the scion and rootstock are brought close together to form a vertical stem. Then, a silicone ring with a diameter of 0.1cm and a height of 0.6cm is used to bind the joint surface and completely seal the cut.

5. The efficient breeding method according to claim 1, characterized in that, In S4, the plant growth chamber was equipped with a 16h / 8h light-dark cycle, with a constant light intensity of 120µmol / m². 2 •s, with a constant temperature of 23℃, the scion will sprout new shoots after growing in the plant growth chamber for 2 months, indicating that the scion has survived.

6. The efficient breeding method according to claim 1, characterized in that, In step S1, the method for propagating seedlings in the nursery is as follows: Step 1: Seedbed preparation During land preparation, apply 5000-10000 kg of well-rotted organic fertilizer and 20-25 kg of superphosphate per mu as base fertilizer. 5-7 days before sowing, spray the seedbed surface with 0.4-0.5% potassium permanganate for soil disinfection. Mix 2-3 g of phoxim with an appropriate amount of fine soil per square meter for soil pest control. When constructing the seedbed, ensure that the seedbed surface is 10-20 cm higher than the walkway and leave drainage ditches between the seedbed surfaces. After the seedbed is formed, harrow and level it, and water it thoroughly before sowing. Step 2: Sowing and Seedling Raising Sowing time is after the soil thaws in spring, when the average daily temperature is stable at 11-14℃ and the temperature at a vertical depth of 10cm is stable at 7-10℃. 5-7 days before sowing, choose a sunny day, spread the cleaned winter-stored seeds evenly on the plastic sheet or canvas inside a temporary plastic greenhouse, with a seed layer thickness of 5-10cm. Keep the seed moisture at 55-65%. At night or on cloudy days, pile the seeds up and spread them out during the day, turning them over in time to prevent rotting. When at least 30% of the seeds show white sprouts and ≥50% of the seeds have turned their tips, it is ready for sowing. The sowing method is as follows: sow in furrows with a sowing width of 9-12cm and a furrow depth of 0.4-0.7cm, cover with soil with a thickness of 0.3-0.5cm, compact with a manual roller after covering with soil, spray water to replenish the moisture of the seed layer, and add more soil to cover the seeds that are exposed after spraying water. Step 3: Seedling Management Seedlings emerge 15-20 days after sowing. During the emergence period, water 3-5 times a day, with each watering amount being 40-50 mL / m². Watering should be done before 9:00 AM and after 4:00 PM. Increase the watering frequency by 1-2 times during windy or hot weather. After emergence, spray 2-3 times a day, increasing the water amount to 80-100 mL / m² each time to encourage root growth.

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