Method for rapid propagation of maesa japonica by cuttage

Abstract

The present application relates to the technical field of plant asexual reproduction, and particularly discloses a cutting propagation method for Lindleya benzoe. The method comprises the following steps: pretreating the base of cuttings with an activating solution containing oligosaccharide signal molecules and active components of root exudates of Lindleya benzoe, then cutting the cuttings in a special induction matrix containing slow-release plant growth regulators, and performing programmed induction of dark culture and light culture; during the light culture stage, dilute solution of the activating solution is applied to the base of the cuttings as an auxiliary inducer. The present application integrates endogenous signal activation, slow-release continuous supply and phased synergistic induction, systematically simulates and strengthens the chemical biological microenvironment of natural rooting, effectively activates the endogenous root potential of the cuttings, and solves the technical problems of low rooting rate, poor root quality and weak transplanting adaptability in conventional methods, thereby realizing efficient and high-quality propagation of Lindleya benzoe cutting seedlings.

Description

Technical Field

[0001] This invention belongs to the field of QQ, specifically a method for rapid propagation of Osmanthus fragrans by cuttings. Background Technology

[0002] In the field of plant propagation, cutting is an important asexual reproduction method for preserving the superior genetic characteristics of the mother plant. Its success hinges on the efficient initiation of adventitious root development and the establishment of a complete root system at the cut site after the cutting detaches from the mother plant. This process is precisely regulated by a complex network of internal physiological and external environmental signals. The polar transport and local accumulation of endogenous hormones (such as auxin) are considered the core driving force inducing root primordia differentiation. Recent research in plant immunity and development has revealed that oligosaccharides with specific structures can act as key molecular signals, activating the expression of defense or development-related genes in plants even at low concentrations. Their mechanism of action is similar to that of "cytokines" in animal cells.

[0003] Although existing technologies widely employ soaking or quick-dip treatment with exogenous plant growth regulators (such as indolebutyric acid) to simulate auxin signals and promote rooting of cuttings, these methods generally have limitations. Treatment with exogenous hormones is often "one-off" and "pulsating," with excessively high initial concentrations easily inhibiting growth or causing abnormal callus proliferation, while the subsequent rapid decay of the concentration fails to provide stable support for continuous root primordia differentiation and elongation. More importantly, this simple external chemical intervention ignores the fact that the cutting is a complete living system, and its rooting process is essentially the orderly expression of its intrinsic genetic program under specific environmental signals. Existing conventional techniques lack the systematic activation of the "rooting potential" within the cutting and fail to simulate the continuous and mild chemo-biological microenvironment mediated by root exudates in natural soil. Therefore, when existing methods are applied to woody ornamental plants that are difficult to root and sensitive to the rooting environment (such as Osmanthus fragrans), they often exhibit technical defects such as unstable rooting rate, poor root quality (few roots, thin and weak roots), long recovery period after transplanting, and large fluctuations in survival rate. This has become a technical bottleneck restricting its large-scale and standardized propagation. Summary of the Invention

[0004] To address the aforementioned technical problems, this invention provides a method for rapid propagation of Osmanthus fragrans cuttings from Hebao Mountain. This method solves the problems of low rooting rate, weak root quality, and unstable transplant survival rate in existing technologies, which are caused by the simple and crude exogenous hormone treatment methods, lack of systematic activation of endogenous rooting potential, and lack of stable rooting microenvironment support.

[0005] A method for propagating Osmanthus fragrans by cuttings includes the following steps:

[0006] S1. Cutting treatment: The base of the Osmanthus fragrans cuttings is immersed in an activation solution for pretreatment. The activation solution contains oligosaccharide signaling molecules and active components derived from the root exudates of Osmanthus fragrans.

[0007] S2, Cuttings: The pretreated cuttings are inserted into an induction substrate, which contains a slow-release induction layer capable of slowly releasing plant growth regulators;

[0008] S3. Cultivation: The cuttings are subjected to dark cultivation and light cultivation in sequence; during the light cultivation stage, the activation solution or a diluted solution of its main active components is applied to the base of the cuttings as an auxiliary inducing agent.

[0009] Preferably, the oligosaccharide signaling molecule is selected from at least one of xyloglucosamine, chitin oligosaccharide, and seaweed oligosaccharide.

[0010] Preferably, the active component derived from the root exudate of Osmanthus fragrans is an organic solvent extract of its root exudate.

[0011] Preferably, the slow-release induction layer comprises a plant growth regulator slow-release unit carried by a biodegradable material; the plant growth regulator includes auxin and brassinolide.

[0012] Preferably, in step S3, the conditions for dark culture are: temperature 23-27℃, relative humidity >85%, no light or weak light, and culture time 7-14 days; the conditions for light culture are: temperature 20-25℃, light intensity 2000-4000 Lux, 10-14 hours of light per day, and relative humidity 75-85%.

[0013] Preferably, the total concentration of oligosaccharide signaling molecules in the activation solution is 10-50 mg / L, and the concentration of the active component of the root exudate on a dry matter basis is 0.05-1.0 mg / L.

[0014] Preferably, the diluent applied in step S3 is the activating solution diluted 20-100 times with sterile water.

[0015] Preferably, the process after step S3 further includes:

[0016] S4. Hardening off and transplanting: After the rooted cuttings have been acclimatized to the environment, they are transplanted into a seedling substrate inoculated with arbuscular mycorrhizal fungi.

[0017] Preferably, the arbuscular mycorrhizal fungus is *Claroideoglomus etunicatum* or *Funneliformis mosseae*.

[0018] Preferably, the process includes the following steps before step S1:

[0019] S0, Pre-strengthening of cuttings: Cuttings taken from the mother plant are hydroponically cultured for a short period of time in a nutrient solution containing trace elements.

[0020] Compared with the prior art, the present invention has the following beneficial effects:

[0021] By combining specific endogenous signal induction, controllable slow-release supply, and phased synergistic stimulation, a complete and efficient regulatory system for the regeneration of Osmanthus fragrans cuttings was constructed. This method not only simulates and enhances the endogenous chemical dialogue and microenvironment required by plants during natural rooting, but also breaks through the technical bottlenecks of conventional cutting techniques, such as difficulty in initiating rooting, uneven rooting quality, and poor adaptability to transplanting, through programmed intervention.

[0022] This method utilizes specific signaling substances derived from the plant itself to activate the endogenous developmental potential of cuttings. Through a two-step strategy of substrate slow release and staged topdressing, it achieves continuous, gentle, and precise guidance of the rooting process, thereby ultimately achieving stable induction of rapid rooting of cuttings, formation of a high-quality root network, and significant improvement in transplant survival rate and seedling resistance. Attached Figure Description

[0023] Figure 1 This is a schematic diagram of the method for propagating Osmanthus fragrans by cuttings according to the present invention. Detailed Implementation

[0024] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0025] like Figure 1 As shown:

[0026] Example 1: A method for propagating Osmanthus fragrans by cuttings, the specific steps of which are as follows:

[0027] S0, pre-strengthening of cuttings:

[0028] Select healthy, semi-lignified branches of Osmanthus fragrans from the current year and cut them into sections about 20 cm long. Immerse the base of the sections in a pre-enhanced nutrient solution, which is a 1 / 4 strength Hoagland nutrient solution with additional 6 µM boric acid and 0.8 µM sodium molybdate. Hydroponically culture for 18 hours at room temperature (about 20-22℃) under natural diffused light.

[0029] S1. Cutting treatment:

[0030] Take out the pre-strengthened branches and make a smooth double-beveled cut 0.5 cm below the node with a sharp blade to make a cutting about 8-10 cm long, retaining the top pair of leaves.

[0031] Preparation of activating solution: Weigh 10.0 mg of xylo-glucosamine oligosaccharide and dissolve it in an appropriate amount of sterile water; add 0.05 mg of ethyl acetate extract powder from the root exudate of Osmanthus fragrans var. pekinensis (preparation method is described below), make up to 1 L, mix well, and use immediately.

[0032] Immerse the base of the cuttings (about 2-3 cm) in the activation solution and let them stand at room temperature (about 25°C) for 60 minutes. Remove them and drain until they no longer drip.

[0033] Note: Preparation of active components from root exudates: Root exudates from healthy Osmanthus fragrans tissue culture seedlings or hydroponic seedlings were collected, filtered through a 0.45 µm filter membrane, and extracted three times with an equal volume of ethyl acetate by shaking. The organic phases were combined and evaporated to dryness at 40°C to obtain a yellowish-brown extract, which was stored at -20°C. Accurate weighing is required before use.

[0034] S2, Cuttings:

[0035] Preparation of the induction matrix: The slow-release induction layer is prepared by mixing decomposed bark, peat moss, and slow-release granules in a volume ratio of 3:1:1. The slow-release granules are prepared as follows: 0.05 mg / L indolebutyric acid (IBA) and 0.01 mg / L brassinolide (BR) are dissolved in water and mixed with 2% sodium alginate solution. The mixture is then dripped into 1% calcium chloride solution to form gel particles. The particles are then coated with a 1% chitosan acetate solution and washed to obtain slow-release granules with a diameter of about 1-2 mm. The upper layer is a breathable and moisture-retaining layer, which is made by mixing vermiculite and perlite in a volume ratio of 1:1.

[0036] Fill the substrate into the acupuncture tray, first fill the lower slow-release induction layer to 2 / 3 of the height of the acupuncture tray, compact it slightly, and then cover it with an upper breathable and moisturizing layer about 2cm thick.

[0037] Make a hole in the center of the substrate with a thin stick, insert the prepared cutting into the hole to a depth of about 3 cm, and gently compact the surrounding substrate.

[0038] S3, Cultivation:

[0039] Dark culture: Place the prepared seedling trays in an artificial climate chamber, set the temperature to 23℃, relative humidity to 90%, and keep it completely dark for 7 days.

[0040] Light culture: After the dark culture is completed, the climate chamber conditions are adjusted to: temperature 20℃, light intensity 2000 Lux (photocycle 10 hours light / 14 hours dark), and relative humidity 75%.

[0041] Assisted induction: On the first day of light culture, dilute the activation solution used in step S1 with sterile water 100 times and gently pour it along the base of the cuttings with a wash bottle until the substrate surface is moist; thereafter, apply it in the same way every 7 days, for a total of 2 applications.

[0042] During the cultivation period, spray an appropriate amount of sterile water according to the substrate humidity to keep the substrate moist but not waterlogged.

[0043] S4. Hardening off and transplanting:

[0044] After about 30 days of light cultivation, when callus tissue has formed at the base of the cuttings and 2-3 new roots with a length of more than 1 cm have grown, hardening off the seedlings can begin.

[0045] Remove the seedling trays from the climate chamber and place them in an acclimatization shed with 50% shade; over 5 days, gradually increase ventilation and reduce the ambient humidity to natural levels (approximately 60%-70%).

[0046] After hardening off the seedlings, carefully remove the cuttings, preserving as much of the original root zone substrate as possible.

[0047] Prepare the seedling substrate: Mix coconut coir, volcanic rock and humus in a volume ratio of 2:1:1, and evenly mix in Claroideoglomus etunicatum spores at a rate of 100 spores per liter of substrate.

[0048] Transplant the cuttings into small nutrient pots containing the seedling substrate, water them thoroughly to help them establish roots, and place them in a semi-shaded environment for routine care.

[0049] Example 2: A method for propagating Osmanthus fragrans by cuttings, the specific steps of which are as follows:

[0050] S0, pre-strengthening of cuttings:

[0051] Select healthy, semi-lignified branches of Osmanthus fragrans from the current year and cut them into sections about 25 cm long. Immerse the base of the sections in a pre-enhanced nutrient solution, which is a 1 / 4 strength Hoagland nutrient solution with additional 7 µM boric acid and 1.0 µM sodium molybdate. Hydroponically culture for 24 hours at room temperature (about 23°C) under natural diffused light.

[0052] S1. Cutting treatment:

[0053] Take out the pre-strengthened branches and make a smooth double-beveled cut 0.8 cm below the node with a sharp blade to make a cutting about 10-12 cm long, retaining 1-2 pairs of leaves at the top.

[0054] Preparation of activating solution: Weigh 15.0 mg of chitosan oligosaccharide (degree of polymerization 5) and dissolve it in an appropriate amount of sterile water; add 0.3 mg of ethyl acetate extract powder from the root secretions of Osmanthus fragrans var. pekinensis, bring the volume to 1 L, mix well, and use immediately.

[0055] Immerse the base of the cutting (about 3 cm) in the activation solution and let it stand at a constant temperature of 25°C for 75 minutes. After removing it, drain it until it stops dripping.

[0056] S2, Cuttings:

[0057] Preparation of the induction matrix: The slow-release induction layer is prepared by mixing decomposed bark, peat moss, and slow-release granules in a volume ratio of 3:1:1. The slow-release granules are prepared as follows: 0.075 mg / L indolebutyric acid (IBA) and 0.015 mg / L brassinolide (BR) are dissolved in water and mixed with 2% sodium alginate solution. The mixture is then dripped into 1% calcium chloride solution to form gel particles. The particles are then coated with a 1% chitosan acetate solution and washed to obtain slow-release granules with a diameter of about 1-2 mm. The upper layer is a breathable and moisture-retaining layer, which is made by mixing vermiculite and perlite in a volume ratio of 1:1.

[0058] Fill the substrate into the apheresis tray, first fill the lower slow-release induction layer to 2 / 3 of the height of the apheresis tray, compact it slightly, and then cover it with an upper breathable and moisturizing layer about 2.5cm thick.

[0059] Make a hole in the center of the substrate with a thin stick, insert the prepared cutting into the hole to a depth of about 3.5 cm, and gently compact the surrounding substrate.

[0060] S3, Cultivation:

[0061] Dark culture: Place the prepared seedling trays in an artificial climate chamber, set the temperature to 25℃, relative humidity to 92%, and keep it completely dark for 10 days.

[0062] Light culture: After the dark culture is completed, the climate chamber conditions are adjusted to: temperature 22℃, light intensity 3000 Lux (photocycle 12 hours light / 12 hours dark), and relative humidity 80%.

[0063] Assisted induction: On the first day of light culture, dilute the activation solution used in step S1 with sterile water 50 times and gently pour it along the base of the cuttings with a wash bottle until the substrate surface is moist; thereafter, apply it in the same way every 5 days for a total of 3 times.

[0064] During the cultivation period, spray an appropriate amount of sterile water according to the substrate humidity to keep the substrate moist but not waterlogged.

[0065] S4. Hardening off and transplanting:

[0066] After about 25 days of light cultivation, when callus tissue has formed at the base of the cuttings and 3-4 new roots with a length of more than 2 cm have grown, hardening off the seedlings can begin.

[0067] Remove the seedling trays from the climate chamber and place them in an acclimatization shed with 60% shade; gradually increase ventilation and reduce ambient humidity to natural levels (approximately 65%) over 6 days.

[0068] After hardening off the seedlings, carefully remove the cuttings, preserving as much of the original root zone substrate as possible.

[0069] Prepare the seedling substrate: Mix coconut coir, volcanic rock and humus in a volume ratio of 2:1:1, and evenly mix in Funneliformis mosseae spores at an inoculation rate of 150 spores per liter of substrate.

[0070] Transplant the cuttings into small nutrient pots containing the seedling substrate, water them thoroughly to help them establish roots, and place them in a semi-shaded environment for routine care.

[0071] Example 3: A method for propagating Osmanthus fragrans by cuttings, the specific steps of which are as follows:

[0072] S0, pre-strengthening of cuttings:

[0073] Select healthy, semi-lignified branches of Osmanthus fragrans from the current year and cut them into sections about 30 cm long. Immerse the base of the sections in a pre-enhanced nutrient solution, which is a 1 / 4 strength Hoagland nutrient solution with additional 8 µM boric acid and 1.2 µM sodium molybdate. Hydroponically culture for 36 hours at room temperature (about 25°C) under natural diffused light.

[0074] S1. Cutting treatment:

[0075] Take out the pre-strengthened branches and make a smooth double-beveled cut 1.0 cm below the node with a sharp blade to make a cutting about 12-15 cm long, retaining the top two pairs of leaves.

[0076] Preparation of activating solution: Weigh 25.0 mg of seaweed oligosaccharide and 25.0 mg of xyloglucosamine oligosaccharide, dissolve them in an appropriate amount of sterile water; add 1.0 mg of ethyl acetate extract powder from the root exudate of Osmanthus fragrans var. sepium, bring the volume to 1 L, mix well, and use immediately.

[0077] Immerse the base of the cutting (approximately 4 cm) in the activation solution and allow it to stand at a constant temperature of 25°C for 90 minutes. Remove the cutting and drain until it no longer drips.

[0078] S2, Cuttings:

[0079] Preparation of the induction matrix: The slow-release induction layer is prepared by mixing decomposed bark, peat moss, and slow-release granules in a volume ratio of 3:1:1. The slow-release granules are prepared as follows: 0.10 mg / L indolebutyric acid (IBA) and 0.02 mg / L brassinolide (BR) are dissolved in water and mixed with 2% sodium alginate solution. The mixture is then dripped into 1% calcium chloride solution to form gel particles. The gel particles are coated with 1% chitosan acetate solution and washed to obtain slow-release granules with a diameter of about 1-2 mm. The upper layer is a breathable and moisture-retaining layer, which is made by mixing vermiculite and perlite in a volume ratio of 1:1.

[0080] Fill the substrate into the acupuncture tray, first fill the lower slow-release induction layer to 2 / 3 of the height of the acupuncture tray, compact it slightly, and then cover it with an upper breathable and moisturizing layer about 3cm thick.

[0081] Make a hole in the center of the substrate with a thin stick, insert the prepared cutting into the hole to a depth of about 4 cm, and gently compact the surrounding substrate.

[0082] S3, Cultivation:

[0083] Dark culture: Place the prepared seedling trays in an artificial climate chamber, set the temperature to 27℃, relative humidity to 95%, and keep it completely dark for 14 days.

[0084] Light culture: After the dark culture is completed, the climate chamber conditions are adjusted to: temperature 25℃, light intensity 4000 Lux (photocycle 14 hours light / 10 hours dark), and relative humidity 85%.

[0085] Assisted induction: On the first day of light culture, dilute the activation solution used in step S1 with sterile water 20 times and gently pour it along the base of the cuttings with a wash bottle until the substrate surface is moist; thereafter, apply it in the same way every 3 days for a total of 4 times.

[0086] During the cultivation period, spray an appropriate amount of sterile water according to the substrate humidity to keep the substrate moist but not waterlogged.

[0087] S4. Hardening off and transplanting:

[0088] After about 20 days of light cultivation, when callus tissue has formed at the base of the cuttings and 4-5 new roots with a length of more than 3 cm have grown, hardening off the seedlings can begin.

[0089] Remove the seedling trays from the climate chamber and place them in an acclimatization shed with 70% shade; gradually increase ventilation and reduce ambient humidity to natural levels (approximately 70%) over 7 days.

[0090] After hardening off the seedlings, carefully remove the cuttings, preserving as much of the original root zone substrate as possible.

[0091] Prepare the seedling substrate: Mix coconut coir, volcanic rock and humus in a volume ratio of 2:1:1, and evenly mix in Claroideoglomus etunicatum spores at a rate of 200 spores per liter of substrate.

[0092] Transplant the cuttings into small nutrient pots containing the seedling substrate, water them thoroughly to help them establish roots, and place them in a semi-shaded environment for routine care.

[0093] Example 4: A method for preparing an active component from the root exudate of Osmanthus fragrans var. pumilus, comprising the following steps:

[0094] Plant preparation: Select uniformly growing Osmanthus fragrans seedlings, carefully wash the soil off the roots, disinfect the surface with 70% ethanol for 30 seconds, then soak in 0.1% HgCl2 solution for 5 minutes, and rinse 5 times with sterile water.

[0095] Pre-culture: Transplant the plants into a sterile hydroponic device containing only basic inorganic salts (such as 1 / 2 strength Hoagland nutrient solution) without organic carbon sources, and pre-culture them for 7 days in an artificial climate chamber (temperature 25±2℃, light 12h / d, humidity 70%) to adapt to the environment and deplete any remaining easily released components in the plant.

[0096] Exudate collection: Replace with fresh inorganic salt culture medium and continue culturing for 14-21 days. Collect all the hydroponic solution during this period, which is the primary solution containing root exudates.

[0097] Preliminary treatment: The collected primary liquid was immediately centrifuged at 8000 rpm for 15 minutes at 4°C. The supernatant was then filtered through 0.45 μm and 0.22 μm microporous membranes for sterilization to obtain a clear root exudate stock solution.

[0098] To concentrate the active substances and remove large amounts of inorganic salts and water, the original solution was extracted and enriched.

[0099] The clarified stock solution was extracted three times with an equal volume of chromatographic grade ethyl acetate in a separatory funnel, with each extraction lasting 10 minutes.

[0100] Combine all ethyl acetate phases (organic phases) and add anhydrous sodium sulfate to remove residual moisture.

[0101] The ethyl acetate phase was concentrated to dryness under reduced pressure using a rotary evaporator in a 40°C water bath to obtain a pale yellow to yellowish-brown viscous or solid extract.

[0102] The extract was reconstituted with analytical grade anhydrous ethanol and diluted to a known volume to prepare a stock solution, which was then stored in a -20°C refrigerator protected from light.

[0103] Before use, dilute to the working concentration with sterile water or appropriate solvent according to the calculated concentration.

[0104] Furthermore, the biodegradable material used in this invention may be one or more of chitosan and its derivatives, natural proteins, and synthetic biodegradable polyesters; the sustained-release unit includes, but is not limited to, microcapsules, gel particles, and porous carrier adsorbent particles prepared using biodegradable materials.

[0105] Furthermore, the auxin used in this invention is preferably indolebutyric acid (IBA), naphthaleneacetic acid (NAA), or their salts; the brassinolide is preferably 24-epibrassinolide or a brassinolide analogue.

[0106] Experimental Example

[0107] I. Experimental Objective

[0108] By setting up a series of comparative ratios, and removing or replacing a certain technical feature, the system evaluates the impact of each feature on the rooting rate of cuttings, root quality, and later growth of plants.

[0109] II. Experimental Materials and Methods

[0110] 1. Experimental Materials

[0111] Plant materials: current-year semi-lignified branches of Osmanthus fragrans from the same mother plant with consistent growth conditions.

[0112] Main reagents and matrix:

[0113] Oligosaccharide: Chitosan oligosaccharide (degree of polymerization 5, biochemical reagent grade)

[0114] Active components of root exudates: Ethyl acetate extract of Osmanthus fragrans root exudates collected and extracted according to the aforementioned standard method.

[0115] Plant growth regulators: Indolebutyric acid (IBA), 24-epibrassinolone (24-epiBL)

[0116] Substrate materials: well-rotted bark, peat moss, vermiculite, and perlite (all commercially available horticultural grade).

[0117] Arbuscular mycorrhizal fungi: Commercial inoculum (containing Funneliformis mosseae spores and hyphae, with ≥80 live spores / gram)

[0118] 2. Experimental Design

[0119] Example 2 was used as the group (T1) of the present invention. The following 5 comparative examples (T2-T6) were set up, each treatment was repeated 3 times, with 10 cuttings per repeat, for a total of 180 cuttings.

[0120] Group Processing description S1 cutting treatment S2 induction matrix S3 light culture-assisted induction T1 (Invention Group) Example 2 Full Solution Activating solution containing chitosan oligosaccharides and root exudate extract Bilayer matrix containing sustained-release particles Apply 50 times diluted activating solution T2 (Comparative Example 1) Only remove the activator solution. Soak only in an equal volume of sterile water Same as T1 Same as T1 T3 (Comparative Example 2) Only the sustained-release matrix was removed Same as T1 Replace with pure vermiculite:perlite (1:1) ordinary matrix Same as T1 T4 (Comparative Example 3) Only remove auxiliary induction Same as T1 Same as T1 Apply only an equal volume of sterile water T5 (Comparative Example 4) Replacement of key components in the activator Activation solution containing only chitosan oligosaccharides (rootless extract) Same as T1 Apply the activating solution diluted solution T6 (Comparative Example 5) Conventional hormone-based rapid application method (control) Dip the sample in 500 mg / L IBA solution for 10 seconds (standard method). Same as T3 (ordinary matrix) none

[0121] 3. Cultivation and Observation

[0122] All cuttings were operated uniformly according to S0 (pre-strengthening), S2 (cutting), S3 (dark / light culture environment parameters) and S4 (hardening and transplanting) of Example 2, with only the specified variables changed according to the table above.

[0123] Observation indicators and time points:

[0124] On the 30th day after cutting: Calculate the rooting rate, measure the average root length, and count the average number of roots.

[0125] 60 days after cutting (30 days after transplanting): Calculate the survival rate, measure the length of new shoots, and determine the relative chlorophyll content (SPAD value) of leaves.

[0126] 60 days after transplanting: Sampling was used to determine the mycorrhizal infection rate (comparison between T1 and T5, T6 only).

[0127] III. Experimental Results

[0128] Table 1: Comparison of rooting status 30 days after cuttings

[0129] Group Rooting rate (%) Average root length (cm) Average number of roots (strips) Root morphology description T1 (This invention) 93.3 5.2 4.7 The root system is robust, with numerous lateral roots and dense root hairs. T2 (Anhydrous Extract Activating Solution) 70.0 3.1 2.8 Root system is thin and weak with few lateral roots T3 (without sustained-release matrix) 76.7 3.8 3.2 The root system is of acceptable length, but its uniformity is poor. T4 (without auxiliary induction) 83.3 4.0 3.5 The root system is well-developed, but the number of lateral roots is slightly less. T5 (without root exudate) 80.0 3.5 3.0 Root system development is average, with few root hairs. T6 (Standard Method) 63.3 2.5 2.1 The root system is short, and occasionally there is swelling of callus tissue.

[0130] Table 2: Plant growth 30 days after transplanting (60 days after cutting)

[0131] Group Survival rate (%) New shoot length (cm) SPAD value T1 (This invention) 96.7 8.5 42.3 T2 76.7 5.1 35.8 T3 83.3 6.0 38.2 T4 90.0 7.2 40.1 T5 86.7 5.8 37.5 T6 73.3 4.3 34.6

[0132] Table 3: Sampling results of mycorrhizal infection rate 60 days after transplanting

[0133] Group Mycorrhizal infection rate (%) Infection description T1 (This invention) 65.2 The plant has abundant arbuscular and vesicular structures, resulting in high infection intensity. T5 (without root exudate) 41.5 The infecting structure is relatively small and the intensity is moderate. T6 (Standard Method) 28.7 Occasional infection points are observed, but their intensity is weak.

[0134] IV. Analysis of Experimental Results

[0135] Synergistic effects of core technological features:

[0136] The entire scheme of this invention (T1) performed best in all observation indicators, with its rooting rate (93.3%), average number of roots (4.7), and later survival rate (96.7%) being significantly higher than all comparative schemes.

[0137] The absence (T2, T3, T4) or replacement (T5) of any single feature leads to a significant decrease in all indicators.

[0138] The key role of active components in root exudates in the "activating solution":

[0139] The T5 group (containing only oligosaccharides and no root exudates) showed significantly lower rooting rate, root length, root number, and later mycorrhizal infection rate compared to the T1 group. This indicates that the active components of root exudates derived from the host itself are the core of the activating solution's "signal transduction" and "specific growth promotion" functions, and their role cannot be replaced by simple oligosaccharide signaling molecules. It may contain specific chemical inducing substances that initiate the rooting process of cuttings and / or chemotactic factors that promote the colonization of beneficial endophytic flora.

[0140] The sustained effects of the "slow-release induction matrix":

[0141] Although the rooting rate of group T3 (using ordinary substrate) was acceptable in the early stage (30 days), its root quality (average number of roots, uniformity) and later growth (new shoot length) were significantly lower than those of group T1. This indicates that the stable and continuous hormone and nutrient microenvironment provided by the slow-release induction layer is crucial for the establishment of root quality and the later growth of the plant, avoiding the problems of excessively high initial hormone concentration or insufficient subsequent supply in conventional methods.

[0142] The consolidation and strengthening effect of "light culture-assisted induction":

[0143] The rooting rate of group T4 (without auxiliary induction) was not significantly different from that of group T1, but its average number of roots and new shoot growth were significantly reduced. This indicates that the main function of reusing the activating solution component (S3) during the light culture stage is not to initiate rooting, but to consolidate and strengthen the rooting process, promote the development of lateral roots and facilitate the smooth transition of the plant to autotrophic growth, thereby improving the quality of seedlings.

[0144] Advantages compared to traditional methods:

[0145] The conventional hormone-based rapid dipping method (T6) showed the lowest results across all indicators. Compared to the conventional method, the method of this invention increases the rooting rate by approximately 30 percentage points, more than doubles the average number of roots, and produces healthy root morphology, avoiding abnormal callus tissue caused by improper hormone treatment. This fully demonstrates the advanced nature and practicality of the invention.

[0146] The embodiments of the present invention are given for the purposes of illustration and description. Although embodiments of the present invention have been shown and described above, it is understood that the above embodiments are exemplary and should not be construed as limiting the present invention. Any changes, modifications, substitutions and variations made by those skilled in the art to the above embodiments within the scope of the present invention should be included within the protection scope of the present invention.

Claims

1. A method for propagating Osmanthus fragrans by cuttings, characterized in that, Includes the following steps: S1. Cutting treatment: The base of the Osmanthus fragrans cuttings is immersed in an activation solution for pretreatment. The activation solution contains oligosaccharide signaling molecules and active components derived from the root exudates of Osmanthus fragrans. S2, Cuttings: The pretreated cuttings are inserted into an induction substrate, which contains a slow-release induction layer capable of slowly releasing plant growth regulators; S3. Cultivation: The cuttings are subjected to dark cultivation and light cultivation in sequence; during the light cultivation stage, the activation solution or a diluted solution of its main active components is applied to the base of the cuttings as an auxiliary inducing agent.

2. The method for propagating Osmanthus fragrans by cuttings according to claim 1, characterized in that, The oligosaccharide signaling molecule is selected from at least one of xyloglucosamine, chitin oligosaccharide, and seaweed oligosaccharide.

3. The method for propagating Osmanthus fragrans by cuttings according to claim 1 or 2, characterized in that, The active component derived from the root exudates of Osmanthus fragrans var. pumilum is an organic solvent extract of its root exudates.

4. The method for propagating Osmanthus fragrans by cuttings according to claim 1, characterized in that, The slow-release induction layer contains a plant growth regulator slow-release unit carried by a biodegradable material; the plant growth regulator includes auxin and brassinolide.

5. The method for propagating Osmanthus fragrans by cuttings according to claim 1, characterized in that, In step S3, the conditions for dark culture are: temperature 23-27℃, relative humidity >85%, no light or weak light, and culture time 7-14 days; the conditions for light culture are: temperature 20-25℃, light intensity 2000-4000 Lux, 10-14 hours of light per day, and relative humidity 75-85%.

6. The method for propagating Osmanthus fragrans by cuttings according to claim 1, characterized in that, The total concentration of oligosaccharide signaling molecules in the activation solution is 10-50 mg / L, and the concentration of the active component of the root exudate is 0.05-1.0 mg / L on a dry matter basis.

7. The method for propagating Osmanthus fragrans by cuttings according to claim 1, characterized in that, The diluent applied in step S3 is the activating solution diluted 20-100 times with sterile water.

8. The method for propagating Osmanthus fragrans by cuttings according to claim 1, characterized in that, The S3 step is followed by: S4. Hardening off and transplanting: After the rooted cuttings have been acclimatized to the environment, they are transplanted into a seedling substrate inoculated with arbuscular mycorrhizal fungi.

9. The method for propagating Osmanthus fragrans by cuttings according to claim 8, characterized in that, The arbuscular mycorrhizal fungi mentioned are *Claroideoglomus etunicatum* or *Funneliformismosseae*.

10. The method for propagating Osmanthus fragrans by cuttings according to claim 1, characterized in that, The steps preceding step S1 also include: S0, Pre-strengthening of cuttings: Cuttings taken from the mother plant are hydroponically cultured for a short period of time in a nutrient solution containing trace elements.

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