Method for improving sowing and germination of ilex pubescens

Abstract

The invention belongs to the technical field of cultivation of ilex pubescens, and relates to a method for improving sowing and germination of ilex pubescens, which specifically comprises the following steps: step 1, seed selection; 2, lysozyme is used for removing fruit membranous layers, and the fruits and a compound enzyme solution are mixed and subjected to stack retting; 3, healthy seeds are screened; step 4, coating and refrigerating; 5, mixing the seeds with the sowing substrate for germination; according to the method, through multiple means of enzymolysis targeted barrier breaking, healthy seed flotation, coating stress-resistant refrigeration, bionic matrix sowing and the like, the germination period of the ilex rosea seeds is remarkably shortened, the germination rate of the ilex rosea seeds is remarkably increased, and the problems that a traditional sowing method of the ilex rosea seeds is low in germination rate and long in germination time are thoroughly solved.

Description

Technical Field

[0001] This invention belongs to the technical field of Luofu holly cultivation, and relates to a method for improving the germination rate of Luofu holly seeds. Background Technology

[0002] As a rare and valuable garden tree species, the seeds of *Ilex tutcheri* are difficult to germinate due to their deep dormancy and unique pericarp structure, which is a core bottleneck restricting large-scale cultivation. Conventional sowing methods suffer from the following defects, making germination of *Ilex tutcheri* seeds extremely difficult, and even when germination occurs, the germination rate is very low and the germination time is long:

[0003] 1) The dormancy mechanism has not been effectively broken. *Ilex laurentii* seeds exhibit dual dormancy characteristics (physical barrier + physiological inhibitory substances). Traditional methods (such as low-temperature sand stratification and gibberellin soaking) target only one factor, resulting in insufficient germination rate and a long germination cycle. Furthermore, the pericarp hinders water and gas exchange in the seeds, and conventional sulfuric acid corrosion or mechanical shell breaking methods easily damage the embryo, leading to seed rot. 2) The fruit processing is crude. Current technology directly peels the seeds, ignoring the influence of residual inhibitors (such as pectin and phenols) in the pulp. Incompletely removed pulp continues to release germination inhibitors, reducing seed viability. Float separation relies solely on water density sorting, failing to accurately remove empty, shriveled, or diseased seeds, resulting in a low selection rate of healthy seeds. 3) Stratification germination efficiency is low. Traditional cold storage (0-5℃) uses a single temperature and humidity level, failing to simulate the temperature spectrum changes during natural overwintering of seeds. This leads to insufficient embryo ripening, poor germination uniformity, and the lack of seed coating protection, making seeds susceptible to water loss or fungal infection during long-term cold storage, resulting in a high rate of mold. 4) Poor adaptability to the sowing environment. The sowing substrate is mostly of a single component, with an imbalance between water retention and air permeability. Seeds are prone to oxygen deficiency and root rot or insufficient water. The shading rate is not controlled. Strong light inhibits the elongation of the radicle, while weak light delays the shedding of the cotyledons, resulting in a low germination rate.

[0004] Therefore, providing a method to improve the germination rate of Ilex laurentii seeds has become an urgent problem to be solved. Summary of the Invention

[0005] To address the aforementioned problems, this invention proposes a method for improving the germination rate of Ilex laurentii seeds, specifically including the following steps:

[0006] Step 1: From December to January of the following year, when the average daily temperature is 10-16℃, select mature fruits of Ilex laurentii that are bright red in color and free from disease and insect spots.

[0007] Step 2: Spray the lysozyme solution onto the surface of the Ilex laurentii fruit at a rate of 50-60 mL / kg of fruit. Let it stand for 2-3 hours to degrade the surface film. Then, mix the Ilex laurentii fruit with the compound enzyme solution and compost at 25-30℃ and 70-80% RH for 3-5 days, turning it over every 1-2 days during the composting process. After composting is complete, the pulp will liquefy to obtain Ilex laurentii seeds.

[0008] Preferably, the lysozyme solution contains 2000-3000 U / g of lysozyme based on water.

[0009] Preferably, the mass ratio of the *Ilex laurentii* fruit to the compound enzyme solution is 100:(0.5-1). Most preferably, the compound enzyme solution, based on water, comprises 3000-4000 U / g cellulase, 2500-3500 U / g ligninase, 2000-3000 U / g pectinase, and 1500-2500 U / g xylanase.

[0010] Step 3: After the composting is completed, wash the Ilex laurentii seeds with clean water and remove the soft and rotten pericarps. Use flotation to screen the healthy Ilex laurentii seeds. Drain the healthy Ilex laurentii seeds and spread them horizontally on a mesh screen. Air dry them at 18-22℃ and 45-50% RH until the seed moisture content is 18-20%.

[0011] Preferably, the flotation method involves mixing Ilex laurentii seeds with a sodium chloride solution of 2-3% by mass at a mass ratio of 1:3, stirring at 90-120 rpm for 20-30 seconds, then letting it stand for 1.5-2 minutes to remove the floating seeds and retain the seeds that sink to the bottom. The seeds that sink to the bottom are then removed and washed with clean water.

[0012] Step 4: After air-drying, spray the seed surface with a coating agent at a rate of 20-30 mL / kg of seeds. Store the coated seeds at 3-4℃ and 65-70% RH for 4-8 days.

[0013] Preferably, the coating agent comprises sodium alginate, nano-silica, gibberellin, proline, and water in a mass ratio of 2:0.5:0.005:0.01:100.

[0014] Step 5: After refrigeration, the seeds are sown and germinated in a seedling device. A 4-6cm thick layer of sowing substrate is laid at the bottom of the seedling device. The seeds of Ilex laurentii are evenly sown on the sowing substrate, and then covered with a 0.5-1cm thick layer of sowing substrate. The seeds are cultured at 20-30℃, 60-70% RH, and 70-75% shading until germination.

[0015] Preferably, the moisture content of the sowing substrate is 70-75%. Most preferably, the sowing substrate comprises peat moss, vermiculite, pine needle soil, earthworm castings and potassium dihydrogen phosphate in a mass ratio of (10-12):(2-3):(3-4):(2-4):(0.01-0.02).

[0016] The present invention has the following advantages:

[0017] (1) Systematically breaking the dormancy mechanism: Lysozyme and a complex enzyme work together to degrade the membranous exocarp and the fleshy mesocarp (commonly known as the pulp). Lysozyme precisely degrades the exocarp, breaking through physical barriers, while the four enzymes (cellulase + ligninase + pectinase + xylanase) liquefy the pulp, thoroughly removing residual inhibitors and avoiding chemical dormancy. Compared to traditional acid etching methods that damage the pericarp structure, the enzymatic hydrolysis method of this invention only targets and decomposes obstacles, preserving the integrity of the embryo.

[0018] (2) Refined seed processing procedure: Sodium chloride solution is used to screen seeds, and density difference is used to efficiently separate healthy seeds (seeds that sink to the bottom) and eliminate empty and shriveled seeds. The screening rate of healthy seeds is significantly improved. Seeds are dried in the shade at low temperature and low humidity to avoid seed dehydration damage and maintain cell membrane stability.

[0019] (3) Stratification can inhibit metabolism, activate embryo maturation, simulate the natural overwintering temperature spectrum, and significantly improve the germination rate of embryos. The coating agent contains gibberellin (to break physiological dormancy), proline (to resist cold stress), and nano-silica (to prevent mold), which significantly reduces the mold rate of seeds during cold storage.

[0020] (4) The biomimetic sowing environment is constructed. The sowing substrate provides 70-75% water content and slow-release nutrients, maintains 70-75% water content and is breathable and prevents compaction. The 70-75% shading rate matches the native environment under the Luofu holly forest, avoids light inhibition, and significantly improves the cotyledon molting synchronization rate. Detailed Implementation

[0021] The technical solutions in the embodiments of the invention are described clearly and completely below. 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.

[0022] Example 1

[0023] Materials preparation:

[0024] The complex enzyme solution comprises, based on water, 3500 U / g cellulase, 3000 U / g ligninase, 2500 U / g pectinase, and 2000 U / g xylanase.

[0025] Coating agent: sodium alginate, nano silica, gibberellin, proline and water, in a mass ratio of 2:0.5:0.005:0.01:100.

[0026] Sowing substrate: peat moss, vermiculite, pine needle soil, earthworm castings and potassium dihydrogen phosphate, in a mass ratio of 11:2.5:3.5:3:0.015.

[0027] Specific implementation steps:

[0028] Step 1: From December to January of the following year, when the average daily temperature is 10-16℃, select mature fruits of Ilex laurentii that are bright red in color and free from disease and insect spots.

[0029] Step 2: Spray a 2500 U / g lysozyme solution onto the surface of the Ilex laurentii fruit at a rate of 55 mL / kg of fruit. Let it stand for 2.5 hours to degrade the surface film. Then, mix the Ilex laurentii fruit and the compound enzyme solution at a mass ratio of 100:1 and compost at 28℃ and 75% RH for 4 days, turning it over once a day during the composting process. After composting is complete, the pulp will liquefy to obtain Ilex laurentii seeds.

[0030] Step 3: After composting, wash the Ilex laurentii seeds with clean water and remove any soft or rotten pericarps. Mix the Ilex laurentii seeds with a 2% sodium chloride solution at a mass ratio of 1:3, stir at 100 rpm for 20 seconds, and then let stand for 2 minutes. Remove the floating seeds and keep the ones that sink to the bottom. After removing the sinking seeds, wash them with clean water. Drain the healthy Ilex laurentii seeds and spread them horizontally on a mesh screen. Then, air dry them at 20°C and 48% RH until the seed moisture content is 18-20%.

[0031] Step 4: After air-drying, spray the seed surface with a coating agent at a rate of 25 mL / kg of seeds, and then refrigerate the coated seeds at 3.5℃ and 63% for 6 days.

[0032] Step 5: After refrigeration, the seeds are sown and germinated in a seedling device. A 5cm thick sowing substrate is laid at the bottom of the seedling device, and the seeds of Ilex laurentii are sown on the sowing substrate. Then, the seeds of Ilex laurentii are covered with a 1cm thick sowing substrate. The seeds are cultured at 25℃, 65% RH, and 75% shading until germination. The moisture content of the sowing substrate is 75%.

[0033] Experimental Example 1

[0034] Experimental group: The seeds of Ilex laurentii were sown and germinated according to the method in Example 1.

[0035] Control group: Refer to the cultivation methods of Ilex laurentii in Volume 1 of "Complete Guide to Ex-situ Cultivated Plants in China" (Aquifoliaceae).

[0036] Sample size: 300 seeds per group, 3 replicates.

[0037] Observation indicators: germination rate, germination cycle, healthy seed screening rate, mold rate, and cotyledon dehulling rate.

[0038] Table 1 Comparison of experimental data (120 days after sowing)

[0039] experimental group control group Improvement effect Germination rate (%) 90.3±2.1 42.7±3.8 ↑111.5％ Germination initiation time (days) 23.5 ± 1.2 days 98.7±5.3 ↓73.8％ Germination uniformity (number of days for 80% of seeds to germinate) 28-35 days 95-110 days Time reduced by 70% Healthy seed screening rate (%) 92.5±1.5 65.3±4.2 ↑41.7％ Refrigerated mold rate (%) 3.2±0.8 37.6±3.1 ↓91.5％ Cotyledon molting success rate (%) 94.1±1.7 58.9±3.5 ↑59.8％

[0040] As shown in Table 1, the method of this invention achieves a breakthrough improvement in germination performance, reaching 90.3% in the experimental group and only 42.7% in the control group, an improvement of 111.5%, proving that enzymatic hydrolysis and segmented cold storage completely break seed dormancy. The germination cycle is shortened by 73.8%, with the experimental group initiating germination in 23.5 days, compared to 89.7 days in the control group. 80% of the seeds germinate within 28-35 days, compared to 95-110 days in the control group, meeting the time requirements for large-scale seedling cultivation. The healthy seed screening rate is increased by 41.7%, with the sodium chloride flotation method achieving an effective seed retention rate of 92.5%, compared to 65.3% in the control group. 98.7% of the floating seeds are empty / damaged, significantly reducing ineffective sowing. The cold storage mold rate is reduced by 91.5%, with the coating agent controlling the mold rate to 3.2%, compared to 37.6% in the control group, solving the problem of long-term cold storage contamination. The cotyledon molting success rate was increased by 59.8%. The biomimetic substrate with 70-75% shading and layered water control resulted in a molting rate of 94.1%, compared to 58.9% in the control group, ensuring the uniformity of seedlings.

[0041] The above description of the disclosed embodiments enables those skilled in the art to make or use the invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the invention. Therefore, the invention is not to be limited to the embodiments shown herein, but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A method for improving the germination rate of Ilex laurentii seeds, characterized in that, Includes the following steps: Step 1: Select ripe fruits of Ilex laurentii that are bright red in color and free from disease and insect spots; Step 2: Spray the lysozyme solution onto the surface of the Ilex laurentii fruit at a rate of 50-60 mL / kg of fruit. Let it stand. Mix the Ilex laurentii fruit with the compound enzyme solution and compost at 25-30℃ and 70-80% RH for 3-5 days, turning it over once every 1-2 days during this period. Step 3: After the composting is completed, wash the Ilex laurentii seeds with clean water and remove the soft and rotten pericarps. Use the flotation method to screen the healthy Ilex laurentii seeds. Drain the healthy Ilex laurentii seeds and spread them horizontally on a mesh screen. Air dry them at 18-22℃ and 45-50% RH until the seed moisture content is 18-20%. Step 4: After air-drying, spray the seed surface with a coating agent and store the coated seeds at 3-4℃ and 65-70% RH for 4-8 days. Step 5: Sow and germinate the refrigerated seeds. First, lay out the sowing substrate, then sow the Ilex laurentii seeds on the sowing substrate, and then cover the Ilex laurentii seeds with the sowing substrate. Cultivate at 20-30℃, 60-70% RH, and 70-75% shading until germination.

2. The method for improving the germination rate of Ilex laurentii seeds according to claim 1, characterized in that, The lysozyme solution described in step two contains 2000-3000 U / g of lysozyme based on water.

3. The method for improving the germination rate of Ilex pubescens seeds according to claim 1, characterized in that, In step two, the mass ratio of Ilex laurentii fruit to the compound enzyme solution is 100:(0.5-1).

4. The method for improving the germination rate of Ilex pubescens seeds according to claim 1, characterized in that, The complex enzyme solution described in step two, based on water, includes 3000-4000 U / g of cellulase, 2500-3500 U / g of ligninase, 2000-3000 U / g of pectinase, and 1500-2500 U / g of xylanase.

5. The method for improving the germination rate of Ilex laurentii seeds according to claim 1, characterized in that, The flotation method described in step three involves mixing Ilex laurentii seeds with a sodium chloride solution of 2-3% by mass at a mass ratio of 1:3, stirring at 90-120 rpm for 20-30 seconds, and then letting it stand for 1.5-2 minutes to remove the floating seeds and retain the seeds that sink to the bottom. The seeds that sink to the bottom are then removed and washed with clean water.

6. The method for improving the germination rate of Ilex laurentii seeds according to claim 1, characterized in that, The coating agent mentioned in step four includes sodium alginate, nano silica, gibberellin, proline and water, in a mass ratio of 2:0.5:0.005:0.01:

100.

7. The method for improving the germination rate of Ilex laurentii seeds according to claim 1, characterized in that, The sowing substrate mentioned in step five includes peat moss, vermiculite, pine needle soil, earthworm castings and potassium dihydrogen phosphate, in a mass ratio of (10-12):(2-3):(3-4):(2-4):(0.01-0.02).

8. The method for improving the germination rate of Ilex laurentii seeds according to claim 1, characterized in that, The moisture content of the sowing substrate mentioned in step five is 70-75%.