A method for seedling raising by sowing of an olea europaea seed

Abstract

This invention discloses a method for sowing and raising olive seeds, belonging to the field of seed propagation technology. It involves using unripe olives as raw material, crushing the shell with a vise to significantly improve the kernel perfection rate; then, enzymatic hydrolysis of the kernel epidermis using protease to reduce the chance of mold growth due to prolonged low-temperature stratification, greatly increasing the germination rate; next, using perlite as a substrate instead of sand for low-temperature stratification of the olive seeds, making it easier to control the substrate moisture content and reducing seed mold; finally, sowing germinated seeds by burying them in the soil to avoid mold rot caused by prolonged contact with the cultivation substrate. Using this method, the germination rate of olive seeds can be significantly increased from the current 40%-60% to 80%-90%, while the cultivation time is significantly shortened from 4-6 months to 2-3 months, with more uniform germination time, vigorous seedling growth, and well-developed root systems.

Description

Technical Field

[0001] This invention belongs to the field of breeding technology, specifically relating to a rapid and efficient method for sowing and raising olive seeds. Background Technology

[0002] Olive oil, a high-grade edible oil, boasts numerous health benefits and is favored by many Chinese consumers, with its annual consumption showing rapid growth. However, premature aging remains a persistent problem plaguing the entire olive oil industry. Some scholars argue that the large number of seedlings currently needed for production are primarily obtained through cuttings of high-yielding trees. However, the roots of these cuttings originate from callus tissue, making them prone to rotting in moist soil. Furthermore, because cuttings lack a taproot, their root systems are relatively weak and shallow, typically only 60-80cm deep, easily leading to premature aging of the tree. In contrast, seedlings propagated from olive seeds possess a distinct taproot and a more developed root system, with the taproot penetrating more than 1 meter into the soil. They also have a longer economic lifespan, vigorous growth, and significant advantages in disease resistance, cold resistance, and drought resistance. Therefore, it is generally believed that grafting high-yielding varieties onto seedling rootstock offers significant advantages over the current direct cutting method and is more effective in addressing the problem of premature aging in olive trees. In addition, scientific research also needs to rely on the large-scale and rapid propagation of olive seedlings to carry out the breeding of new olive varieties.

[0003] The commonly used outdoor stratification method for seedling cultivation in production and research involves mixing seeds with moist sand in a specific ratio. The moisture content of the sand should be controlled at around 3% (too low moisture content hinders seed germination, while too high moisture content leads to extensive mold growth). The sand bed needs to be turned over every week to improve ventilation and prevent mold. After four months of stratification, the olive seeds begin to germinate after breaking dormancy due to the low temperatures of autumn and winter, typically taking six months to a year to complete the emergence of seedlings. This method is not only time-consuming and results in inconsistent emergence times, making large-scale propagation difficult in the short term and hindering the widespread use of seedlings, but also generally results in a low germination rate, usually only 40%–70%. While common plant growth regulators, such as gibberellin and 6-benzylaminopurine, have a significant effect on seed enlargement, they are not very effective in improving seed germination rate or shortening germination time. Therefore, developing new technologies for olive seedling cultivation from seed is of positive significance for promoting the development of the olive industry. Summary of the Invention

[0004] To address the shortcomings of existing olive seed propagation techniques, this invention provides a rapid and efficient method for sowing and raising olive seeds.

[0005] To achieve the above objectives, the present invention adopts the following technical solution:

[0006] A method for sowing and raising seedlings from olive seeds, comprising the following steps:

[0007] (1) Seed collection and storage

[0008] After crushing the unripe fresh olives, collect the seeds, add clean river sand and rub and stir to remove the pulp and oil attached to the seed shell. Then pick out the seeds from the sand, wash them clean with water, dry them in a ventilated place, pack them in a cloth bag and hang them in a dry and ventilated room for long-term storage.

[0009] This invention uses unripe olive fruit as breeding material. The kernels are plump, avoiding the shriveling caused by incomplete seed development and preventing deep dormancy and splitting of the seeds due to overripe fruit, thus improving seed germination rate. Furthermore, the drying process of fresh olive seeds widens the gap between the kernel and the hard shell, significantly reducing kernel breakage during subsequent mechanical shell cracking.

[0010] (2) Seed breaking, flotation, enzymatic hydrolysis, disinfection, soaking, stratification and germination.

[0011] ① Shell breaking and flotation

[0012] After crushing the preserved olive seeds with pliers, carefully break open the seed shell, remove the kernel, and place it in 30wt% salt water for flotation. After skimming off the empty and shriveled kernels that float to the surface, wash the kernels that sink with clean water to remove the salt water residue on the surface.

[0013] This invention utilizes vises as the shell-breaking tool, which significantly improves the success rate of shell-breaking kernels. Compared to existing methods of shell-breaking using hammers, pliers, or shears, or by corroding with concentrated sulfuric acid, the germination rate using vises is typically higher than 80%, reaching up to 95%, demonstrating a clear advantage.

[0014] ② Enzymatic hydrolysis of seed coat, disinfection, and seed soaking

[0015] The floated kernels were placed in a constant-temperature magnetic stirring pot, and enzyme solution was added until it just covered the seeds. Enzymatic hydrolysis was carried out by stirring at a constant temperature to remove protein components on the seed coat and enhance seed coat permeability. After enzymatic hydrolysis, the kernels were filtered out and washed three times with clean water to remove residual protease adhering to the kernel surface. Then, they were first soaked in 0.1 vol% carbendazim solution for 0.5 hours for disinfection, and then soaked in 1000 mg / L gibberellin solution for 4 hours. The enzymatic hydrolysis used neutral protease with a concentration of 0.1~0.01 U / mL, a hydrolysis time of 0.5~2 h, and a hydrolysis temperature of 40~50℃.

[0016] Compared to surgical cutting or treatment with acids, alkalis, or cellulase, enzymatic hydrolysis of the seed coat with proteases improves seed coat permeability, thus promoting kernel germination, and causes less damage to the seed coat structure. Furthermore, removing surface proteins reduces the likelihood of mold growth in the kernel, significantly decreasing seed rot caused by microbial invasion during subsequent low-temperature stratification.

[0017] ③ Stratification for germination

[0018] Mix the soaked seed kernels with perlite at a ratio of 1 seed to 1 mL of perlite, place the mixture in a seedling tray, and water thoroughly until water flows out from the bottom of the tray. Then, place the tray in a constant temperature incubator at 10-15℃ in the dark to promote germination. Every 7 days, remove the seedling tray and rinse the perlite in the tray with a 750 mg / kg chlorothalonil solution (the volume of chlorothalonil solution should be 3 times the volume of perlite). Replenish the water needed for seed germination to reduce the number of microorganisms in the substrate and prevent mold growth. Every 10 days, turn the perlite in the tray and remove any moldy seeds to promote ventilation and aeration of the substrate. After 20 days of stratification, the seeds will begin to germinate. At this time, select seeds with a radicle length of more than 2 mm and sow them. After 30 days of stratification, most seeds will have germinated. Stop stratification and discard any ungerminated seeds. The washed perlite substrate can be reused for the next seed germination.

[0019] This invention uses perlite as the stratification material for low-temperature stratification, which allows for easy moisture control and avoids the seed mold problems caused by poor moisture content and aeration in sand stratification, a method often used in seedling cultivation with the seed coat intact. Furthermore, stratifying olive kernels at 10-15℃ effectively inhibits mold growth and ensures a rapid dormancy period. The perlite substrate used for stratification is also recyclable, resulting in low costs.

[0020] (3) Sowing and seedling raising

[0021] Soak compressed seedling blocks (3×4cm in size, mainly composed of peat) in water until softened. Use a hole opener to make a hole about 1 cm deep in the upper substrate of each seedling block. Take a pre-germinated seed kernel, place it root-down in the hole of the seedling block, one seed per hole. Fill the gap between the seed kernel and the hole with 20-mesh perlite granules to complete the sowing. Then, transfer the seedling blocks to a plant light incubator for cultivation. The cultivation temperature is 20℃~25℃, the humidity is 80%~90%, and the illuminance is 2000~4000 LUX. Leaves will begin to unfold after 1 week. After 2 weeks, most of the seeds will have basically unfolded leaves. After 3 weeks, most of them will have grown 4 true leaves.

[0022] Given that olive trees thrive in dry climates and are intolerant of dampness, and that their seeds are highly susceptible to water absorption and rotting, this invention employs a root-burying method for sowing. This method involves placing only the root tip of the germinated seed in contact with the moist substrate during sowing, while keeping the stems and leaves dry. This method helps prevent seed rot and achieves a germination rate of over 95%.

[0023] (4) Hardening off seedlings and transplanting

[0024] Mix peat and perlite in a 1:1 (v / v) ratio to form a substrate. Then, take a seedling block with 4 true leaves and bury it in the substrate of the seedling bag. Transplant it into a small greenhouse for cultivation. During the day, lift the plastic film on both sides of the greenhouse to allow ventilation and harden the seedlings. At night, cover the plastic film tightly. After one week, move the seedling bag outdoors. After growing outdoors for one year, when the ground diameter of the olive seedlings reaches 5mm, they can be used as grafting rootstock for olive cultivation or for breeding new olive varieties.

[0025] The green-ripe fresh olives used in step (1) refer to olives that have just entered the ripening stage, that is, olives whose peels are turning from green to light green.

[0026] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0027] This invention employs several novel methods for sowing and cultivating olive seeds, including using a vise to break the seed coat, enzymatic hydrolysis of the seed coat with protease, low-temperature stratification with perlite, and root-burying sowing. The vise-based method significantly improves the kernel's perfection rate by squeezing and breaking the seed coat. Then, enzymatic hydrolysis of the kernel skin reduces the likelihood of mold growth due to prolonged low-temperature stratification, greatly increasing the germination rate. Perlite is used instead of sand for low-temperature stratification, making it easier to control substrate moisture content and reducing seed mold. Finally, germinating seeds are sown using root-burying sowing to prevent mold rot caused by prolonged contact with the cultivation substrate. Testing shows that this method increases the germination rate of olive seeds from the current 40%-70% to 80%-90%, shortens the cultivation time from 4-6 months to 2-3 months, results in more consistent germination, and produces seedlings with vigorous growth and well-developed root systems. Attached Figure Description

[0028] Figure 1 This is a comparative graph showing the effects of different enzymatic hydrolysis methods on the germination rate of olive seeds. 1-Control, 2-Cellulase, 3-Hemicellulase, 4-Pectinase, 5-Amylase, 6-Lipase, 7-Ligninase, 8-Mixed enzyme (equal amounts of 2-7), 9-Protein; the amount of enzyme used was 0.1% (w / w) of the seed weight.

[0029] Figure 2 This is a comparative graph showing the effects of different stratified substrates on the germination rate of enzymatically hydrolyzed olive seeds.

[0030] Figure 3 This is a comparative graph showing the effect of perlite dosage on the germination rate of enzymatically hydrolyzed olive seeds.

[0031] Figure 4 This is a comparative diagram showing the effects of different sowing methods on the germination rate of seeds. The methods are as follows: 1- Seeds with roots facing down are placed in the holes of the seedling block and filled with perlite; 2- Sowing in a sand bed and covering with 2cm of sand; 3- Sowing in a soil bed and covering with 2cm of soil; 4- Sowing in a peat bed and covering with 2cm of peat; 5- Sowing in a coconut coir bed and covering with 2cm of coconut coir; 6- Seeds with roots facing down are inserted into the surface of a sand bed; 7- Seeds with roots facing down are inserted into the surface of a soil bed; 8- Seeds with roots facing down are inserted into the surface of a peat bed; 9- Seeds with roots facing down are inserted into the surface of a coconut coir bed. Detailed Implementation

[0032] A method for sowing and raising seedlings from olive seeds, comprising the following steps:

[0033] (1) Seed collection and storage

[0034] After crushing the unripe fresh olives, collect the seeds, add clean river sand and rub and stir to remove the pulp and oil attached to the seed shell. Then pick out the seeds from the sand, wash them clean with water, dry them in a ventilated place, pack them in a cloth bag and hang them in a dry and ventilated room for long-term storage.

[0035] (2) Seed breaking, flotation, enzymatic hydrolysis, disinfection, soaking, stratification and germination.

[0036] ① Shell breaking and flotation

[0037] After crushing the preserved olive seeds with pliers, carefully break open the seed shell, remove the kernel, and place it in 30wt% salt water for flotation. After skimming off the empty and shriveled kernels that float to the surface, wash the kernels that sink with clean water to remove the salt water residue on the surface.

[0038] ② Enzymatic hydrolysis of seed coat, disinfection, and seed soaking

[0039] The floated kernels were placed in a constant-temperature magnetic stirring pot, and enzyme solution was added until it just covered the seeds. Enzymatic hydrolysis was carried out by stirring at a constant temperature to remove protein components on the seed coat and enhance seed coat permeability. After enzymatic hydrolysis, the kernels were filtered out and washed three times with clean water to remove residual protease adhering to the kernel surface. Then, they were first soaked in 0.1 vol% carbendazim solution for 0.5 hours for disinfection, and then soaked in 1000 mg / L gibberellin solution for 4 hours. The enzymatic hydrolysis used neutral protease with a concentration of 0.1~0.01 U / mL, a hydrolysis time of 0.5~2 h, and a hydrolysis temperature of 40~50℃.

[0040] ③ Stratification for germination

[0041] Mix the soaked seed kernels with perlite at a ratio of 1 seed to 1 mL of perlite, place the mixture in a seedling tray, and water thoroughly until water flows out from the bottom of the tray. Then, place the tray in a constant temperature incubator at 10-15℃ in the dark to promote germination. Every 7 days, remove the seedling tray and rinse the perlite in the tray with a 750 mg / kg chlorothalonil solution (the volume of chlorothalonil solution should be 3 times the volume of perlite). Replenish the water needed for seed germination to reduce the number of microorganisms in the substrate and prevent mold growth. Every 10 days, turn the perlite in the tray and remove any moldy seeds to promote ventilation and aeration of the substrate. After 20 days of stratification, the seeds will begin to germinate. At this time, select seeds with a radicle length of more than 2 mm and sow them. After 30 days of stratification, most seeds will have germinated. Stop stratification and discard any ungerminated seeds. The washed perlite substrate can be reused for the next seed germination.

[0042] (3) Sowing and seedling raising

[0043] Soak the compressed seedling blocks (3×4cm in size, mainly composed of peat) in water until they expand. Use a hole opener to make a hole about 1 cm deep in the upper part of the substrate of each seedling block. Take the pre-germinated kernels and place them root-down into the holes of the seedling blocks, one kernel per hole. Fill the gaps between the kernels and the holes with 20-mesh perlite granules to complete the sowing. Then, transfer the seedling blocks to a plant light incubator for cultivation. The cultivation temperature is 20℃~25℃, the humidity is 80%~90%, and the illuminance is 2000~4000 LUX. After 1 week, the leaves will begin to unfold. After 2 weeks, all the seeds will have basically finished unfolding their leaves. After 3 weeks, most of them will have grown 4 true leaves.

[0044] (4) Hardening off seedlings and transplanting

[0045] Mix peat and perlite in a 1:1 (v / v) ratio to form a substrate. Then, take a seedling block with 4 true leaves and bury it in the substrate of the seedling bag. Transplant it into a small greenhouse for cultivation. During the day, lift the plastic film on both sides of the greenhouse to allow ventilation and harden the seedlings. At night, cover the plastic film tightly. After one week, move the seedling bag outdoors. After growing outdoors for one year, when the ground diameter of the olive seedlings reaches 5mm, they can be used as grafting rootstock for olive cultivation or for breeding new olive varieties.

[0046] The green-ripe fresh olives used in step (1) refer to olives that have just entered the ripening stage, that is, olives whose peels are turning from green to light green.

[0047] Figure 1This is a comparative graph showing the effects of different enzymatic hydrolysis methods on the germination rate of olive seeds. 1-Control, 2-Cellulase, 3-Hemicellulase, 4-Pectinase, 5-Amylase, 6-Lipase, 7-Ligninase, 8-Mixed enzyme (equal parts of 2-7), 9-Protein; the enzyme dosage for all methods was 0.1% (w / w) of the seed weight. Figure 1 The results show that enzymatic hydrolysis of olive seeds using proteases significantly improves the germination rate compared to hydrolysis using other single or combined enzymes. This is likely because enzymatic hydrolysis removes surface proteins from the seeds, increases the carbon-to-nitrogen ratio, inhibits microbial growth, and reduces the likelihood of mold growth before germination, thereby increasing the germination rate.

[0048] Figure 2 This is a comparative graph showing the effects of different stratification substrates on the germination rate of enzymatically hydrolyzed olive seeds. Figure 2 The results show that using perlite stratification to germinate olive seeds results in a higher germination rate than other common substrates. This is likely because perlite has low water absorption, does not easily accumulate water, has good aeration, and is less likely to cause seed mold.

[0049] Figure 3 This is a comparative graph showing the effect of perlite dosage on the germination rate of enzymatically hydrolyzed olive seeds. Figure 3 The results show that adding more than 1 mL of perlite to each seed can achieve a higher seed germination rate; while adding 0.5 mL of perlite to each seed may lead to seed suffocation due to insufficient oxygen supply from the substrate pores, thus reducing the germination rate.

[0050] Figure 4 This is a comparative diagram showing the impact of different sowing methods on the germination rate of seeds. The methods are as follows: 1- Insert seeds root-down into the holes of the seedling block and fill with perlite; 2- Sow seeds on a sand bed and cover with 2cm of sand; 3- Sow seeds on a soil bed and cover with 2cm of soil; 4- Sow seeds on a peat bed and cover with 2cm of peat; 5- Sow seeds on a coconut coir bed and cover with 2cm of coconut coir; 6- Insert seeds root-down into the holes of the seedling block and fill with sand; 7- Insert seeds root-down onto the surface of a sand bed; 8- Insert seeds root-down onto the surface of a soil bed; 9- Insert seeds root-down onto the surface of a peat bed. Figure 4 The results showed that Group 1 had the highest germination rate, which may be attributed to the fact that the seedling block substrate was made of pressed peat, which is not easy to loosen after being perforated, making it easier to complete this special sowing operation than other substrates; and while the seed roots penetrate the substrate to obtain nutrients, the perlite filling can effectively prevent the kernel surface from contacting the moist substrate, thus preventing seed mold.

[0051] To make the content of this invention easier to understand, the technical solution of this invention will be further described below with reference to specific embodiments, but this invention is not limited thereto.

[0052] Example 1

[0053] 1. Seed collection and storage

[0054] The unripe fresh olives of the Ezhi No. 8 variety were placed on a wire mesh with a 3-mesh aperture. Wearing rain boots, the fruit was stomped and rubbed vigorously to break up the pulp and sieve it, leaving the seeds on the mesh. The seeds were collected, rubbed and stirred with clean river sand to further remove the pulp and oil attached to the seed shell. The seeds were then picked out from the sand, thoroughly washed in clean water, placed in a ventilated place to air dry, packed in cloth bags, and hung in a dry and ventilated indoor place for long-term storage.

[0055] 2. Seed breaking, flotation, enzymatic hydrolysis, disinfection, soaking, and stratification for germination.

[0056] (1) Shell breaking and flotation

[0057] Dry and preserved olive seeds were held horizontally between commercially available Type 50 vises. The vises were slowly rotated to tighten the handle, gradually increasing the pressure on the seeds until the seed coat cracked. The vises were then immediately released, and the cracked seed coat was carefully broken open. The kernels were removed and placed in a 30wt% salt solution for flotation. Empty and shriveled kernels that floated to the surface were removed, and the intact kernels that sank to the bottom were kept. The remaining salt solution on the surface of the kernels was washed away with clean water. The rate of superior seeds was 75%.

[0058] (2) Enzymatic hydrolysis of seed coat, disinfection, and seed soaking

[0059] After flotation, the high-quality seed kernels were placed in a beaker, a magnetic stir bar was added, and the beaker was placed in a constant temperature magnetic stirring pot. 0.1 U / mL of neutral protease enzyme solution was added until the seeds were just submerged. The mixture was stirred at 50℃ for 0.5 h to remove protein components on the seed coat and enhance seed coat permeability. After the enzymatic hydrolysis was completed, the seed kernels were filtered out and washed three times with clean water to remove residual protease adhering to the seed kernel surface. The seed kernels were then first soaked in 0.1 vol% carbendazim solution for 0.5 h for disinfection, and then soaked in 1000 mg / L gibberellin solution for 4 h.

[0060] (3) Stratification for germination

[0061] Mix the soaked seed kernels with perlite at a ratio of 1 seed to 1 mL of perlite, place the mixture in a seedling tray, and water thoroughly until water flows out from the bottom of the tray. Then, place the tray in a constant temperature incubator at 10-15℃ in the dark to promote germination. Every 7 days, remove the seedling tray and rinse the perlite in the tray with a 750 mg / kg chlorothalonil solution (the volume of chlorothalonil solution should be 3 times the volume of perlite). Replenish the water needed for seed germination to reduce the number of microorganisms in the substrate and prevent mold growth. Every 10 days, turn the perlite in the seedling tray to promote ventilation and remove any moldy seeds. After 20 days of stratification, the seeds will begin to germinate. At this time, select seeds with a radicle length of more than 2 mm and sow them. After 30 days of stratification, most of the seeds will have germinated. Stop stratification and discard any ungerminated seeds.

[0062] 3. Sowing and seedling raising

[0063] First, soak the compressed seedling blocks (3×4cm in size, mainly composed of peat) in tap water until they expand. Then, use a hole opener to make a hole about 1 cm deep in the substrate on the top of the seedling block. Take a pre-germinated olive kernel, place it root-down in the hole of the seedling block, one kernel per hole, and fill the gap between the kernel and the hole with 20-mesh perlite granules to complete the sowing. Then, transfer the seedling blocks to a plant light incubator for cultivation at a temperature of 25℃, a humidity of 80%, and an illuminance of 4000 LUX. Leaves will begin to unfold after one week, and after two weeks, most seeds will have basically completed leaf unfolding. After three weeks, most seeds will have grown four true leaves.

[0064] 4. Hardening off seedlings and transplanting

[0065] Mix peat and perlite in a 1:1 (v / v) ratio to form a substrate. Then, take seedling blocks with four true leaves, bury them in the substrate in seedling bags, and move them into small greenhouses for cultivation. During the day, lift the plastic film on both sides of the greenhouse to allow for ventilation and hardening off the seedlings, and cover the film tightly at night. After one week, move the seedling bags outdoors. From the start of germination to obtaining seedlings, the required time is two months, and the seedling survival rate (based on superior varieties) is 82%. After one year, the ground diameter of the olive seedlings reaches about 5 mm, which can be used as grafting rootstock for olive planting, or for breeding new varieties.

[0066] Example 2

[0067] 1. Seed collection and storage

[0068] Place the ripe, green olives of the As variety on a 7-mesh wire mesh, put on rain boots, and vigorously stomp and rub the fruit to break up the pulp and sieve it, leaving the seeds on the mesh. Collect the seeds, add clean river sand, and rub and stir to further remove the pulp and oil attached to the seed shell. Pick out the seeds from the sand, wash them thoroughly in clean water, place them in a ventilated place to air dry, pack them in a cloth bag, and hang them in a dry, well-ventilated room for long-term storage.

[0069] 2. Seed breaking, flotation, enzymatic hydrolysis, disinfection, soaking, and stratification for germination.

[0070] (1) Shell breaking and flotation

[0071] Dry and preserved olive seeds were held horizontally between commercially available Type 50 vises. The vises were slowly rotated to tighten the handle, gradually increasing the pressure on the seeds until the seed coat cracked. The vises were then immediately released, and the cracked seed coat was carefully broken open. The kernels were removed and placed in a 30wt% salt solution for flotation. Empty and shriveled kernels that floated to the surface were removed, and the intact kernels that sank to the bottom were kept. The remaining salt solution on the surface of the kernels was washed away with clean water. The rate of superior seeds was measured to be 80%.

[0072] (2) Enzymatic hydrolysis of seed coat, disinfection, and seed soaking

[0073] After flotation, the high-quality seed kernels were placed in a beaker, a magnetic stir bar was added, and the beaker was placed in a constant temperature magnetic stirring pot. 0.05 U / mL of neutral protease enzyme solution was added until the seeds were just submerged. The mixture was stirred at 40℃ for 1 hour to remove protein components on the seed coat and enhance seed coat permeability. After the enzymatic hydrolysis was completed, the seed kernels were filtered out and washed three times with clean water to remove residual protease adhering to the seed kernel surface. The seed kernels were then first soaked in 0.1 vol% carbendazim solution for 0.5 hours for disinfection, and then soaked in 1000 mg / L gibberellin solution for 4 hours.

[0074] (3) Stratification for germination

[0075] Mix the soaked seed kernels with perlite at a ratio of 1 seed to 1 mL of perlite, place the mixture in a seedling tray, and water thoroughly until water flows out from the bottom of the tray. Then, transfer the tray to a constant temperature incubator at 10-15℃ in the dark to promote germination. Every 7 days, remove the seedling tray and rinse the perlite in the tray with a 750 mg / kg chlorothalonil solution (the volume of chlorothalonil solution should be 3 times the volume of perlite). Replenish the water needed for seed germination to reduce the number of microorganisms in the substrate and prevent mold growth. Every 10 days, turn the perlite in the tray to promote ventilation and remove any moldy seeds. After 20 days of stratification, the seeds will begin to germinate. At this time, select seeds with a radicle length of more than 2 mm and sow them. After 30 days of stratification, most seeds will have germinated. Stop stratification and discard any ungerminated seeds.

[0076] 3. Sowing and seedling raising

[0077] First, soak the compressed seedling blocks (3×4cm in size, mainly composed of peat) in tap water until they expand. Then, use a hole opener to make a hole about 1 cm deep in the substrate on the top of the seedling block. Take a pre-germinated olive kernel, place it root-down in the hole of the seedling block, one kernel per hole, and fill the gap between the kernel and the hole with 20-mesh perlite granules to complete the sowing. Then, transfer the seedling blocks to a plant light incubator for cultivation at a temperature of 25℃, a humidity of 90%, and an illuminance of 2000 LUX. Leaves will begin to unfold after 1 week, and after 2 weeks, most seeds will have basically completed leaf unfolding. After 3 weeks, most seeds will have grown 4 true leaves.

[0078] 4. Hardening off seedlings and transplanting

[0079] Prepare a substrate by mixing peat and perlite in a 1:1 (v / v) ratio. Then, take seedling blocks with four true leaves, bury them in the substrate in seedling bags, and move them into small greenhouses for cultivation. During the day, open the greenhouse film on both sides to allow ventilation and harden the seedlings, and cover the film tightly at night. After one week, move the seedling bags outdoors. The required seedling time from the start of bud break is 75 days, and the seedling survival rate (based on superior varieties) is 85%. After one year, the ground diameter of the olive seedlings reaches about 5 mm, which can be used as grafting rootstock for cultivation or for the breeding of new olive varieties.

[0080] The above description is only a preferred embodiment of the present invention. All equivalent changes and modifications made within the scope of the claims of the present invention should be included in the scope of the present invention.

Claims

1. A method for sowing and raising seedlings of olive seeds, characterized in that, Includes the following steps: (1) Seed collection and storage After crushing the unripe fresh olives, collect the seeds, add clean river sand and rub and stir to remove the pulp and oil attached to the seed shell. Then wash the seeds with clean water, air dry them in a ventilated place, pack them in a cloth bag and store them in a dry and ventilated room. (2) Seed breaking, flotation, enzymatic hydrolysis, disinfection, soaking, stratification and germination. ① Shell breaking and flotation After crushing the preserved olive seeds with pliers, carefully break open the seed shell, remove the kernel, and place it in 30wt% salt water for flotation. After skimming off the empty and shriveled kernels that float to the surface, wash the kernels that sink with clean water to remove the salt water residue on the surface. ② Enzymatic hydrolysis of seed coat, disinfection, and seed soaking Add the floated kernels to the enzyme solution until they just cover the seeds, stir at a constant temperature for enzymatic hydrolysis, filter out the kernels after enzymatic hydrolysis, wash them three times with clean water, first soak them in 0.1 vol% carbendazim solution for 0.5 hours for disinfection, and then soak them in 1000 mg / L gibberellin solution for 4 hours. ③ Stratification for germination After soaking, mix the kernels with perlite in a specific ratio, place the mixture in a seedling tray, and water thoroughly until water flows out from the bottom of the tray. Then, transfer the tray to a constant temperature incubator for low-temperature, light-protected germination. Every 7 days, remove the seedling tray and rinse the perlite with a 750 mg / kg chlorothalonil solution, while replenishing the water needed for seed germination to reduce the substrate bacterial count and prevent mold growth. Every 10 days, turn the perlite in the seedling tray, remove any moldy seeds, and promote ventilation and aeration of the substrate. After 20 days of stratification, the seeds begin to germinate. At this time, select seeds with a radicle length exceeding 2 mm for sowing. After 30 days of stratification, most seeds have germinated. Stop stratification and discard any ungerminated seeds. (3) Sowing and seedling raising Soak the compressed seedling blocks in water until they expand, and use a hole opener to make a hole 1 cm deep in the upper substrate of each seedling block; take the germinated kernels, place them root-down in the holes of the seedling blocks, one kernel per hole, and fill the gaps between the kernels and the holes with 20-mesh perlite granules to complete the sowing; then transfer the seedling blocks into a plant light incubator for cultivation. (4) Hardening off seedlings and transplanting Take a seedling block with 4 true leaves and bury it in the substrate of the seedling bag. Then move it into a small arched greenhouse for cultivation. During the day, lift the plastic film on both sides of the greenhouse to allow for ventilation and hardening off the seedlings. At night, cover the plastic film tightly. After 1 week, move the seedling bag outdoors. Olive seedlings are grown outdoors for one year until their ground diameter reaches 5mm, at which point they can be used as grafting rootstock for olive cultivation or for breeding new olive varieties. The enzymatic hydrolysis in step (2) uses a neutral protease with a concentration of 0.1~0.01 U / mL, a hydrolysis time of 0.5~2h, and a hydrolysis temperature of 40~50℃.

2. The method for sowing and raising seedlings of olive seeds according to claim 1, characterized in that: The unripe fresh olives used in step (1) refer to olives that have just entered the ripening stage.

3. The method for sowing and raising seedlings of olive seeds according to claim 1, characterized in that: In step (2), the seeds are mixed with 1 mL of perlite at a ratio of 1 seed to 1 mL, and the germination temperature is 10-15℃.

4. The method for sowing and raising seedlings of olive seeds according to claim 1, characterized in that: The main component of the compressed seedling block used in step (3) is peat, and its size is Φ3×4cm.

5. The method for sowing and raising seedlings of olive seeds according to claim 1, characterized in that: The temperature for cultivation in step (3) is 20~25℃, the humidity is 80%~90%, and the illuminance is 2000~4000 LUX.

6. The method for sowing and raising seedlings of olive seeds according to claim 1, characterized in that: The matrix used in step (4) is a mixture of peat and perlite in a volume ratio of 1:1.

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