Method for breeding spores of cibotium barometz and rejuvenating young spores

Abstract

The invention provides a method for breeding spores of cibotium barometz and rejuvenating young spores. The method for artificially cultivating the Cibotium barometz, provided by the invention, comprises the following steps: separately planting sporophyte seedlings of the Cibotium barometz to be rejuvenated into a matrix, and carrying out rejuvenation culture; wherein the substrate is a mixture formed by mixing leaf mold and peat soil according to the volume ratio of (1-1.5): 1 or 1: 1, or a mixture formed by mixing leaf mold, white vermiculite and garden soil according to the volume ratio of 4: 1: 3. The method is low in cost, simple in technology, high in yield, high in seedling growth speed, suitable for large-scale batch production, capable of effectively meeting the rapid breeding requirement of the golden dog and capable of being widely applied to the fields of landscaping, medicinal production, scientific research and the like.

Description

Technical Field

[0001] The invention relates to the technical field of forest seedling cultivation, and in particular to a method for breeding golden dogwood spores and rejuvenating young spore seedlings. Background Art

[0002] Cibotium barometz (Linn.) J.Sm. belongs to the Cibotiaceae family and the Cibotium genus. It is a perennial tall tree-like fern medicinal plant, mainly distributed in tropical and subtropical regions of Asia, especially in southern my country. The rhizome of Cibotium barometz is thick and procumbent, with a cluster of large leaves growing on the top. The petioles are brown and covered with cushion-like golden hairs at the base. It likes scattered light and often grows in a humid and warm forest environment. It is an internationally renowned garden ornamental plant and an important traditional Chinese medicine. Cibotium barometz is famous for its unique appearance and medicinal value. It has an upright plant shape, large and spreading leaves, and a beautiful appearance. It has a very high ornamental value and occupies an important position in horticultural applications. Its rhizome (Chinese medicinal material name "gou ji") is included in the "Chinese Pharmacopoeia" (2020 edition). It has the effects of removing rheumatism, nourishing the liver and kidneys, and strengthening the waist and knees. It is often used to treat rheumatic pain, soreness of the waist and knees, and weakness of the lower limbs. The scales on the rhizome can also be used for external application to stop bleeding. In addition, in recent years, a variety of active ingredients (such as flavonoids and polysaccharide compounds) have been extracted from golden dogs through modern technology, providing a new development direction for pharmaceutical technology, and its health and edible value has gradually been valued.

[0003] Although the golden dog has great ornamental, medicinal and health value, its wild resources are decreasing year by year due to disorderly development. In recent years, due to excessive exploitation of wild golden dog resources and neglect of its growth law and distribution characteristics, the number of wild golden dogs has dropped sharply. At present, the golden dog has been listed in the Convention on International Trade in Endangered Species of Wild Fauna and Flora (Appendix II, CITES2016), and is also listed as a national second-level protected plant in the National Key Protected Wild Plant List (First Batch) . Its endangered status is mainly attributed to its difficulty in reproduction, slow growth, extensive traditional planting methods, and lack of proprietary management technology for ferns. The current planting methods are not ideal in terms of seedling survival rate, seedling growth rate and plant quality, resulting in the development of golden dogs relying more on wild resources, further exacerbating the problem of resource scarcity. With the increasing demand for golden dog resources in the fields of horticulture, medicine and scientific research, it has become a top priority to protect wild golden dog resources and develop effective breeding methods.

[0004] The reproduction of Cibotium chinense mainly includes three methods: sexual reproduction (spore reproduction), asexual reproduction (ramet reproduction) and in vitro reproduction (tissue culture). Among them, spore reproduction often leads to long seedling raising cycle, low survival rate, poor adaptability of seedlings and even large-scale seedling death due to problems such as low germination rate, uneven emergence, slow growth of seedlings and high proportion of weak seedlings. These problems seriously restrict the promotion and application of artificial large-scale breeding of Cibotium chinense and become the key technical bottleneck that needs to be solved urgently. Summary of the invention

[0005] In order to solve the problem of the shortage of golden dog resources, the inventor of the present invention has obtained a set of methods for artificial breeding of golden dog after a lot of exploration, which has a high survival rate and short time consumption. The method of the present invention aims at the problems of golden dog seedling cultivation such as low spore germination rate, uneven emergence, and yellow leaves of gametophyte and sporophyte, and proposes a solution with the characteristics of uniform emergence, rapid emergence, and strong growth.

[0006] The method for artificially cultivating Cibotium barometz provided by the present invention comprises the following steps: transplanting Cibotium barometz sporophyte seedlings to be rejuvenated into a matrix for rejuvenation culture; wherein the matrix is ​​a mixture of leaf mold and peat soil in a volume ratio of (1-1.5):1 or 1:1, or a mixture of leaf mold, white vermiculite and garden soil in a volume ratio of 4:1:3.

[0007] Among them, leaf humus, also known as humus soil, is a nutrient soil formed by the decomposition and fermentation of plant branches and leaves in the soil by microorganisms. As a specific example, the organic matter content in leaf humus is 45.7% and the soil bulk density is 0.36g / cm 3 , PH (water immersion) 6.0, the texture of leaf mold is black, and it contains organic soil particles.

[0008] Among them, peat soil refers to the soil in some river and lake sedimentary low plains and mountain valleys, where due to long-term water accumulation, dense aquatic vegetation, and in the absence of oxygen, a large amount of incompletely decomposed plant residues accumulate and form a peat layer. As a specific example, the sum of the content of organic matter and humic acid in peat soil is greater than or equal to 50%, porosity: 70-80, PH (water immersion) 5.6-6.5.

[0009] Preferably, in the above method, when the first true leaf of the sporophyte seedling grows to 0.3-0.5 cm, transplanting and rejuvenation culture are carried out.

[0010] Preferably, the rejuvenation culture conditions include: a temperature of 23-25°C, a light intensity of 2500-3500 Lux, a light duration of 12-14 hours or 14 hours per day, and a relative humidity of 65%-85% in the space where the sporophyte seedlings are located. The space where the sporophyte seedlings are located is a space formed by planting the sporophyte seedlings in a seedling container and covering the seedling container with a transparent cover.

[0011] Preferably, the water content of the matrix in the rejuvenation culture is 60% to 65% or 60%.

[0012] Preferably, the rejuvenation culture also includes the step of applying a nutrient solution; specifically, the nutrient solution is a water-soluble fertilizer of a large number of elements; further specifically, the nutrient solution is applied starting from the 90th day after transplanting, and the water-soluble fertilizer of a large number of elements is diluted into an aqueous solution with a concentration of 0.2% (mass percentage concentration), and sprayed once every 7 days, with 200 ml sprayed per 1760 square centimeters each time.

[0013] Among them, macronutrient water-soluble fertilizers are commonly used in plant growth. They are mainly composed of three essential macronutrients for plants: nitrogen (N), phosphorus (P) and potassium (K). These elements play a key role in the growth and development of plants, and because they can be completely dissolved in water and are easy for plants to absorb, they are widely used in agricultural production, such as drip irrigation, spraying and foliar fertilization. The ratio we use is a balanced type, which contains 20% nitrogen (N), 20% phosphorus (P2O5) and 20% potassium (K2O) (model: 20-20-20+TE, implementation standard NY / T 1107-2020).

[0014] Preferably, the method further comprises a step of hardening the seedlings, wherein the sporophyte seedlings to be hardened are planted separately, and the substrate is a mixture of leaf mold and peat soil in a volume ratio of (1-1.5):1 or 1:1.

[0015] Preferably, the sporophyte seedlings to be hardened are sporophyte seedlings with a height of 8.5-10.0 cm or sporophyte seedlings each having 5 to 8 leaves.

[0016] Preferably, the conditions for hardening the seedlings include: a temperature of 25-30° C., an air humidity of 60%-65% in the greenhouse, a light intensity of 3000-3700 lux, and a light duration of 12-14 h.

[0017] Preferably, the method comprises the following steps: sowing spores of Cibotium barometz in a substrate, first performing a dark treatment, and then culturing until sporophyte seedlings to be rejuvenated are obtained.

[0018] Preferably, the conditions of the dark treatment include: culturing in the dark for 44 to 48 hours or 48 hours; specifically, the conditions of the dark treatment include: culturing in the dark and at a temperature of 23 to 25°C for 44 to 48 hours or 48 hours; further specifically, the conditions of the dark treatment include: culturing in the dark, at an initial substrate humidity of 60% or 60% to 65%, at a temperature of 23 to 25°C, and in a closed condition for 44 to 48 hours or 48 hours; further specifically, the conditions of the dark treatment include: culturing in the dark, at an initial substrate humidity of 60% or 60% to 65%, at a temperature of 23 to 25°C, and in a closed condition for 44 to 48 hours or 48 hours, without replenishing water during the period. The closed means that a transparent cover is placed on the seedling container, and then the gap between the container and the cover is sealed to form a closed environment space, and the spores or sporophyte seedlings are in the seedling container.

[0019] Preferably, the step of re-culturing to obtain the sporophyte seedlings to be rejuvenated includes the step of culturing until the sexual organs of the gametophyte mature under the following conditions: the ambient temperature is 23-25°C, the light intensity is 2500-3500 Lux, the light is on for 12-14 hours or 14 hours a day, and the relative humidity of the air in the environment where the gametophyte is located is 65-85%. Specifically, the gametophyte sexual organs are cultured until they mature under the following conditions: the ambient temperature is 23-25°C, the light intensity is 2500-3500 Lux, the light is on for 12-14 hours or 14 hours a day, the relative humidity of the air in the environment where the gametophyte is located is 65-85%, and it is airtight. The relative humidity of the air in the environment where the gametophyte is located is the relative humidity of the air in the seedling container covered with a transparent cover. Airtight refers to the space environment formed by covering the seedling container with a transparent cover and sealing the gap between the container and the cover. If the relative humidity of the air is not within the range of 65-85%, water is added to make the relative humidity of the air within the range of 65-85%. After the water is added, the airtight state is immediately restored.

[0020] Preferably, after the gametocyte sexual organs mature, the following steps are included:

[0021] (1) The relative humidity of the gametophyte environment is adjusted to 65-85%, and then cultured for 10 days under airtight conditions at a constant temperature of 25°C without adding water during the period; specifically, the relative humidity of the gametophyte environment is adjusted to 65-85%, and then cultured for 10 days under airtight conditions at a constant temperature of 25°C, with a light intensity of 2500-3500 Lux, and 12-14 hours or 14 hours of light per day, without adding water during the period; wherein the relative humidity of the gametophyte environment is the relative humidity of the air in the seedling container covered with a transparent cover. The airtight refers to the space environment formed by covering the seedling container with a transparent cover and sealing the gap between the container and the cover.

[0022] (2) The sealed condition is then released, and the gametophytes are placed in a temperature range of 23-25° C., a light intensity of 2500-3500 Lux, a light intensity of 12-14 hours or 14 hours per day, and a relative humidity of 22-25% for 22-24 hours or 24 hours; wherein the relative humidity of 22-25% is the condition in a greenhouse, and after the sealed condition is released, the gametophytes are exposed to the greenhouse, and the relative humidity of the greenhouse is 22-25%. The sealed condition is released, i.e., the transparent cover is removed.

[0023] (3) Promote fertilization by spraying water mist.

[0024] Preferably, the method comprises the following steps: after fertilization, culturing until the sporophyte seedlings germinate; more specifically, after fertilization, culturing until the sporophyte seedlings germinate under the following conditions: ambient temperature of 23-25°C, light intensity of 2500-3500 Lux, 12-14 hours or 14 hours of light per day, relative air humidity of 65-85%; more specifically, the culturing is carried out under closed conditions. Wherein closed refers to the space environment formed by covering the seedling container with a transparent cover and sealing the gap between the container and the cover.

[0025] Preferably, after the sporophyte seedlings germinate, they are cultured under the following conditions until the sporophyte seedlings to be rejuvenated are obtained: light intensity of 3000-4000 Lux, 12-14 hours or 14 hours of light per day, temperature of 26-27° C., and relative air humidity maintained at 55%-70%; wherein the relative air humidity is the relative air humidity in the seedling raising container covered with a transparent cover.

[0026] Preferably, the substrate for spore inoculation is peat soil, specifically, Pinnaceous peat soil, with a particle size of 0-6 mm and a pH value of 5.5-6.0.

[0027] Preferably, the initial moisture content of the substrate used for spore inoculation is 60%.

[0028] Herein, sporophyte seedlings and young sporelings have the same meaning and are interchangeable.

[0029] The golden dog seedling raising method provided by the present invention can realize rapid batch seedling raising of spores, significantly improve the germination rate of spores (increased to 3000 plants / tray, i.e., 3000 plants / 0.176 square meters), the uniformity of emergence is high, the gametophyte and sporophyte are strong, and the method also shortens the seedling growth period to 120 days. In short, the method has low cost, simple technology, high yield, fast seedling growth rate, and is suitable for large-scale batch production. It can effectively meet the rapid breeding needs of golden dogs and can be widely used in the fields of landscaping, medicinal production, and scientific research. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 It is the spore leaf of Golden Dogwood.

[0031] Figure 2 It is the mature gametophyte of golden retriever.

[0032] Figure 3 It is the spore seedling of Golden Dog.

[0033] Figure 4 It is the medium for transplanting and rejuvenation with different treatments.

[0034] Figure 5 This is the seedling acclimatization situation of T1 group on the 15th day.

[0035] Figure 6 Shown are the growth and development of young sporelings in different groups on the 45th day.

[0036] Figure 7 This is the growth and development of seedlings in group T1 on the 90th day.

[0037] Figure 8 This is the growth and development of seedlings in group T1 on the 120th day.

[0038] Fig. 9 This is the growth and development of golden retriever seedlings on the 150th day. DETAILED DESCRIPTION

[0039] 1. Collection of golden dog spores

[0040] Select the spore leaves of golden dogwood with a spore sorium cracking rate of 20% to 30%, and mature and well-growing plants for collection. When collecting, put the spore leaves into a rectangular paper bag made of 2 to 3 layers of newspaper, seal it and place it in a cool and ventilated place to dry naturally for 10 to 15 days. Before sowing, gently lift the dried paper bag, cut off a corner, and the spores fall on the white paper after tapping. If the spores are mixed with impurities, spread them evenly, tilt the white paper and flick it to separate the spores from the impurities. After repeating the process until the impurities are completely removed, collect the purified spores into a centrifuge tube and store them in a refrigerator at 4°C to ensure the activity and quality of the spores.

[0041] 2. Inoculation of Golden Dog Spores

[0042] The soil used for sowing is Danish imported Pins peat soil (particle size 0-6mm, pH 5.5-6.0) provided by Beijing Zhonghong Lianke Biotechnology Co., Ltd. After the peat soil is wrapped with newspaper, it is sterilized by high-pressure steam at 125℃ for 25 minutes, and then spread flat in a 55cm×32cm×8cm culture tray after cooling, compacted to a soil layer thickness of 5cm, and sprayed with tap water until the soil moisture content is about 60%. When sowing, 0.01ml of spores are placed at the fold line of the folded A4 paper, and the paper is flicked to evenly spread the spores into the culture tray along the fold line. After sowing, cover the seedling tray with a transparent lid and seal the gap between the tray lids with black electrical tape to keep the substrate moist. No watering is required after sowing. Cover the seedling tray with an opaque black cloth and place it in a culture room with an ambient temperature of 23-25℃ for 48 hours to promote synchronous germination of spores and more uniform growth of seedlings. After the above-mentioned dark treatment, the culture environment temperature is controlled at 23-25°C, the light intensity is 2500-3500Lux, and the light is on for 14 hours a day; and the relative humidity of the air (relative humidity of the air in the seedling tray) is maintained at 65%-85% by timely water replenishment, that is, spray water to the matrix in the seedling tray to make the humidity in the seedling tray reach 65-85%, immediately cover the seedling tray with a transparent lid, and seal the gap between the tray lids with electrical tape, and maintain this sealed environment until the following germination begins and then open the lid.

[0043] 3. Artificial Assisted Fertilization and Sporangial Transformation

[0044] 10 to 15 days after sowing, the golden dog spores begin to germinate; about 60 to 80 days later, the gametophyte sexual organs (sperm apparatus and archegonium) mature, and their formation can be observed under a microscope. After the sexual organs mature, open the cover to detect the humidity, spray water mist to keep the humidity at 65 to 85%, and then keep a constant temperature of 25°C, seal the culture plate for 10 days, and no watering is required during this period. During this period, the light intensity is 2500 to 3500 Lux, and the light is 14 hours a day. Subsequently, open the transparent plastic cover, do not spray to maintain humidity, but open the cover to dry naturally in the greenhouse for 24 hours (during this period, the temperature is 23 to 25°C, the light intensity is 2500 to 3500 Lux, the light is 14 hours a day, and the relative humidity of the air in the culture room is 22-25%), to promote the full development of the sperm apparatus and the increase of sperm concentration. After drying, through a small amount of uniform spraying (once in the morning, noon and evening), the sexual organs are fully soaked with water to complete fertilization. After fertilization, the environment is sealed again for 7 days, that is, the cultivation tray is covered with a transparent cover and sealed with electrical tape. During this period, it is not moved or sprayed with water (the relative humidity of the air in the seedling tray is 65-85%) until the sporophyte seedlings germinate. If the seedlings do not grow after 7 days, the transparent cover can be opened for ventilation for 3 days, and the spraying and sealing steps are repeated. During the entire fertilization process, the temperature is 23-25°C, the light intensity is 2500-3500 Lux, and the light is 14 hours a day. After the seedlings grow out, the light intensity is increased to 3000-4000 Lux, the light is 14 hours a day, the room temperature is controlled at 26-27°C, and the relative humidity of the air (relative humidity of the air in the seedling tray) is maintained at 55%-70%. After this treatment, the germination rate of golden dog spores can reach 3000 plants / tray (3000 plants / 0.176 square meters).

[0045] 4. Seedling rejuvenation

[0046] After the golden dog spore seedlings grow out, observe the growth every 3 days. When the first true leaf grows to 0.3-0.5 cm, use tweezers to transplant the seedlings to the culture tray of the same specifications as the sowing for division and rejuvenation. The study found that the subculture matrix has a significant effect on the rejuvenation of golden dog spore seedlings. In order to screen the best matrix, the experiment used leaf mold and peat soil as the main materials, coconut bricks, white vermiculite (5-8mm), pond mud and garden soil as auxiliary materials, and prepared different matrix combinations (specific ratios are shown in Table 1), and the moisture content of the matrix was 60%. The experiment set up 4 treatments (T1-T4), and 3 trays of seedlings were transplanted in each treatment matrix, 100 plants were planted in each tray, and each tray was 1 replicate, with a total of 3 replicates.

[0047] Coconut bricks, white vermiculite, pond mud, and garden soil were purchased from Huihuang Gardening Supplies Store in Longgang District, Shenzhen.

[0048] Leaf humus, also known as humus soil, is a nutrient soil formed by plant branches and leaves in the soil after microbial decomposition and fermentation. In this embodiment, the leaf humus soil was purchased from Shenzhen Green Country Garden Co., Ltd. The organic matter content was 45.7% and the soil bulk density was 0.36g / cm 3 , PH (water immersion) 6.0, the texture of leaf mold is black, and it contains organic soil particles.

[0049] Peat soil: Peat soil refers to soil in some river and lake sedimentary low plains and mountain valleys. Due to long-term accumulation of water, aquatic vegetation is dense, and a large amount of inadequately decomposed plant residues accumulate and form a peat layer under anaerobic conditions. The peat soil in this embodiment was purchased from Changchun Zhuangmiao Peat Technology Co., Ltd., and the product name is the new seedling medium-Jintian brand. In the peat soil, the sum of the content of organic matter and humic acid is greater than or equal to 50%, the porosity is 70-80, and the pH (water immersion) is 5.6-6.5.

[0050] Table 1 Composite seedling medium formula (volume ratio)

[0051]

[0052] The substrate used for division rejuvenation does not need to be sterilized. The division container uses the seedling tray and transparent cover used for sowing, but it is no longer sealed with tape. After transplanting, the culture temperature is controlled at 23-25℃, the light intensity is 2500-3500Lux, and the light time is 14 hours a day. Replenish water in time to keep the relative humidity of the air in the seedling tray at 65%-85%; spray water mist to increase the humidity when the humidity is lower than 65%, and ventilate appropriately when it is higher than 85%. Observe the seedling recovery and the degree of pests and diseases, and record the survival rate (%) of sporophyte seedlings on the 15th, 30th, 45th, 60th, 75th and 90th days after transplanting. The survival rate calculation formula is: number of surviving plants / 100×100%. The plant height was measured on the 30th, 60th and 90th days, and the test results are shown in Table 2.

[0053] Table 2 Growth and development of golden dog spore seedlings under different treatments

[0054]

[0055] Note: Different lowercase letters after the data in the same column indicate significant differences (p < 0.05)

[0056] According to Table 2, different seedling mediums have a significant effect on the growth and development of the golden dog young spore seedlings. The standard for seedling acclimatization is to grow the second true leaf. Only the T1 group medium (leaf humus 1: peat soil 1) reached this standard within 20 days. Its plant height can reach 4.5-5.5 cm within 90 days, showing a good growth state, and the final survival rate is 79.17%. In contrast, the other three groups did not achieve effective seedling acclimatization, and the growth state was poor: the survival rates of the T2 and T3 groups dropped to 4.00% and 4.33% on the 60th day, respectively, and all the seedlings died within 75 days; the survival rate of the T4 group was only 9.67% on the 75th day, and all the seedlings died within 90 days. In addition, the four groups of matrix treatments all had different degrees of algae contamination, among which the T1 group had a lighter degree of contamination, while the other three groups had more serious contamination, which may have an adverse effect on the acclimatization and growth of the seedlings. This experiment showed that the substrate of leaf mold (1): peat soil (1) was most suitable for the transplanting and rejuvenation of young spore seedlings of Golden Dog. In this substrate, the young spore seedlings were able to grow slowly within 20 days, and the plant height could reach 4.5-5.5 cm within 90 days. The survival rate was as high as 79.17%, which was significantly better than other substrate treatment groups.

[0057] 5. Water and fertilizer management

[0058] On the 90th day after transplanting and rejuvenation, start spraying nutrient solution in appropriate amounts to continue cultivation. The nutrient solution uses a large amount of water-soluble fertilizer (implementation standard NY / T 1107-2020), diluted into an aqueous solution with a concentration of 0.2% (mass percentage concentration), sprayed once every 7 days, and 200 ml per tray each time. Under this culture condition, the light intensity is maintained at 2500-3500 Lux, the indoor temperature is maintained at 23-25°C, and the relative humidity of the air (relative humidity of the air in the seedling tray) is maintained at 65%-85%. After spraying nutrient solution for cultivation, within about 120 days, the height of the young spore seedlings of Golden Dog can reach 8.5-10.0 cm, and each seedling can grow 5 to 8 leaves. At this time, the seedlings are growing well, and subsequent potting and hardening can be carried out to promote independent growth and improve adaptability of the plants. When hardening the seedlings, the seedlings with a height of 8.5 to 10.0 cm were planted in pots individually. The hardening medium was selected from the same leaf mold (1) and peat soil (1) as the T1 group. This was the best medium selected in previous experiments and helped to promote the healthy growth of the seedlings. After planting, water was poured in time to fix the roots and placed in a hardening greenhouse. The temperature of the greenhouse was maintained between 25 and 30°C. Water mist was sprayed 2 to 3 times a day, each time for 10 minutes, to ensure that the relative humidity of the culture air was maintained at 60% to 65%. The light intensity for hardening the seedlings was 3000 to 3700 lux, and the light time was 12 to 14 hours. After about 30 days of hardening (about 150 days from the start of spore seedling transplanting), the plant height could reach 12 to 15 cm, and the number of leaves increased to 6 to 9, showing a good growth state and could enter the further cultivation stage.

Claims

1. A method for artificially cultivating Cibotium barometz, comprising the following steps: transplanting the sporophyte seedlings of Cibotium barometz to be rejuvenated into a substrate for rejuvenation culture; wherein: The matrix is ​​a mixture of leaf mold and peat soil in a volume ratio of (1-1.5):1 or 1:1, or a mixture of leaf mold, white vermiculite and garden soil in a volume ratio of 4:1:

3.

2. The method according to claim 1, characterized in that: When the first true leaf of the sporophyte seedling grows to 0.3-0.5 cm, it is transplanted and rejuvenated.

3. The method according to claim 1 or 2, characterized in that: The rejuvenation culture conditions include: a temperature of 23-25° C., a light intensity of 2500-3500 Lux, a light duration of 12-14 hours or 14 hours per day, and a relative humidity of 65%-85% in the space where the sporophyte seedlings are located; and / or, the water content of the substrate in the rejuvenation culture is 60% to 65% or 60%; And / or, the rejuvenation culture also includes the step of applying a nutrient solution; specifically, the nutrient solution is a water-soluble fertilizer of a large number of elements; further specifically, the nutrient solution is applied on the 90th day after transplanting, and the water-soluble fertilizer of a large number of elements is diluted into an aqueous solution with a concentration of 0.2% (mass percentage concentration), and sprayed once every 7 days, with 200 ml sprayed per 1760 square centimeters each time.

4. The method according to any one of claims 1 to 3, characterized in that: The method also includes a seedling hardening step, wherein the sporophyte seedlings to be hardened are planted separately, and the substrate is a mixture of leaf mold and peat soil in a volume ratio of (1-1.5):1 or 1:

1.

5. The method according to any one of claims 1 to 4, characterized in that: The sporophyte seedlings to be hardened are sporophyte seedlings with a height of 8.5-10.0 cm or sporophyte seedlings each having 5 to 8 leaves; And / or, the conditions for hardening the seedlings include: a temperature of 25-30° C., a relative humidity of 60%-65% in the greenhouse, a light intensity of 3000-3700 lux, and a light duration of 12-14 hours.

6. The method according to any one of claims 1 to 5, characterized in that: The method comprises the following steps: sowing spores of Cibotium barometz in a substrate, firstly performing a dark treatment, and then culturing until sporophyte seedlings to be rejuvenated are obtained.

7. The method according to any one of claims 1 to 6, characterized in that: The conditions for the dark treatment include: culturing in the dark for 44 to 48 hours or 48 hours; specifically, the conditions for the dark treatment include: culturing in the dark and at a temperature of 23 to 25°C for 44 to 48 hours or 48 hours; further specifically, the conditions for the dark treatment include: culturing in the dark, with an initial substrate humidity of 60% or 60% to 65%, at a temperature of 23 to 25°C, and in a closed condition for 44 to 48 hours or 48 hours; further specifically, the conditions for the dark treatment include: culturing in the dark, with an initial substrate humidity of 60% or 60% to 65%, at a temperature of 23 to 25°C, and in a closed condition for 44 to 48 hours or 48 hours, without replenishing water during the period.

8. The method according to any one of claims 1 to 7, characterized in that: The step of re-culturing to obtain the sporophyte seedlings to be rejuvenated comprises: culturing until the gametophyte sexual organs mature under the following conditions: the ambient temperature is 23-25°C, the light intensity is 2500-3500 Lux, the light is on for 12-14 hours or 14 hours a day, and the relative humidity of the air in the environment where the gametophyte is located is 65-85%; specifically, the step of re-culturing to obtain the sporophyte seedlings to be rejuvenated comprises: culturing until the gametophyte sexual organs mature under the following conditions: the ambient temperature is 23-25°C, the light intensity is 2500-3500 Lux, the light is on for 12-14 hours or 14 hours a day, and the relative humidity of the air in the environment where the gametophyte is located is 65-85%, and the environment is airtight.

9. The method according to any one of claims 1 to 8, characterized in that: After the gametocyte sexual organs mature, the method comprises the following steps: (1) The relative humidity of the air in the environment where the gametophytes are located is adjusted to 65-85%, and then the gametophytes are cultured in a sealed environment at a constant temperature of 25° C. for 10 days without water supplementation; specifically, the relative humidity of the air in the environment where the gametophytes are located is adjusted to 65-85%, and then the gametophytes are cultured in a sealed environment at a constant temperature of 25° C. with a light intensity of 2500-3500 Lux and a light intensity of 12-14 hours or 14 hours per day for 10 days without water supplementation; (2) the sealed condition is then released, and the gametocytes are placed in a culture medium at 23-25°C, with a light intensity of 2500-3500 Lux, with light exposure for 12-14 hours or 14 hours per day, and a relative humidity of 22-25% for 22-24 hours or 24 hours; (3) Promote fertilization by spraying water mist.

10. The method according to any one of claims 1 to 9, characterized in that: The method comprises the following steps: after fertilization is completed, culturing until the sporophyte seedlings germinate; more specifically, after fertilization is completed, culturing until the sporophyte seedlings germinate under the following conditions: an ambient temperature of 23 to 25°C, a light intensity of 2500 to 3500 Lux, 12 to 14 hours or 14 hours of light per day, and an air relative humidity of 65 to 85%; and even more specifically, the culturing is carried out under closed conditions. and / or, after the sporophyte seedlings germinate, culturing under the following conditions until the sporophyte seedlings to be rejuvenated are obtained: light intensity of 3000-4000 Lux, 12-14 hours or 14 hours of light per day, temperature of 26-27° C., relative air humidity maintained at 55%-70%; And / or, the substrate for spore inoculation is peat soil, specifically Pinnaceous peat soil, with a particle size of 0 to 6 mm and a pH value of 5.5 to 6.0; And / or, the initial moisture content of the substrate used for spore inoculation is 60%.

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