Rapid propagation method for radix ranunculi ternati tissue
By using a multi-component synergistic compounding primary induction medium and a phased temperature, light and humidity control strategy, the problem of balancing rooting rate and root length in the propagation of Cat's Claw Grass was solved, resulting in the cultivation of high-quality and robust seedlings that meet the needs of large-scale production.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2026-01-22
- Publication Date
- 2026-04-07
AI Technical Summary
Existing cat's claw grass propagation techniques cannot simultaneously improve rooting rate and average root length, resulting in poor seedling quality that cannot meet the demand for high-quality and robust seedlings for large-scale production.
The primary induction culture medium, which is synergistically formulated with multiple components, is combined with a gradient-progressive temperature, light, and humidity control strategy to precisely match the physiological rhythm of the entire cycle from seed germination to seedling growth of Cat's Claw. By adding membrane stabilizers, organic nitrogen sources, and natural antioxidant extracts, the needs of bud differentiation and seedling robustness are balanced, and the development of root primordia is promoted.
It achieved a synergistic improvement in both high rooting rate and long root length, cultivating seedlings with robust root systems, and enhancing the survival rate and subsequent growth stability of the seedlings.
Smart Images
Figure CN121795322A_ABST
Abstract
Description
Technical Field
[0001] This invention belongs to the field of plant tissue culture technology, specifically relating to a method for rapid propagation of cat's claw grass tissue. Background Technology
[0002] Cat's Claw Grass (Radix Ranunculi Ternati) is the dried tuberous root of Ranunculata natus Thunb., a plant in the Ranunculaceae family. It possesses properties such as clearing heat and detoxifying, dispersing nodules and reducing swelling, and is an important variety in the Chinese medicinal herb market. However, due to its slow natural propagation rate and limited resources, the supply and demand imbalance of Cat's Claw Grass is becoming increasingly prominent. Therefore, researching an efficient tissue propagation process is of great significance for protecting Cat's Claw Grass resources and meeting market demand.
[0003] However, existing cat's claw plant propagation techniques either focus on increasing the propagation coefficient or only on optimizing the efficiency of a single step, failing to solve the core problem of the difficulty in simultaneously achieving a high rooting rate and average root length. Either the pursuit of a high rooting rate results in a shorter average root length, affecting transplant survival rate and subsequent growth stability; or attempts to increase root length are accompanied by a significant decrease in rooting rate, making it impossible to cultivate high-quality seedlings with multiple and robust roots. This technological bottleneck directly restricts the improvement of seedling quality, making it difficult to meet the demand for high-quality, robust seedlings in large-scale production.
[0004] Therefore, developing a rapid tissue propagation method for cat's claw grass that can simultaneously improve rooting rate and average root length and cultivate robust root seedlings has significant practical application value. Summary of the Invention
[0005] The purpose of this invention is to provide a method for rapid propagation of cat's claw grass tissue.
[0006] To achieve the above objectives, the present invention provides the following technical solution: A method for rapid propagation of cat's claw plant tissue includes the following steps: (1) Seed selection and disinfection: Select viable seeds, wash the seeds, soak them in mercuric chloride, disinfect them with alcohol, and finally rinse them with sterile water. (2) Preparation of primary induction medium: Preparation of primary induction medium; (3) Seed induction and proliferation: The sterilized seeds were inoculated on the primary induction medium and cultured. After 4-6 weeks of culture, the seeds germinated into seedlings. The seedlings were then transferred to the subculture induction medium for proliferation culture, and subcultured every 3-4 weeks. (4) Rooting culture: Transfer the propagated seedlings to the rooting culture medium and culture for 2-3 weeks. Adventitious buds can take root and form complete plants. (5) Hardening off and transplanting: After the roots have developed, the complete plants are moved to the greenhouse for hardening off. After hardening off for 1-2 weeks, the plants are transplanted into peat soil.
[0007] Preferably, the primary induction medium is based on MS medium and contains the following components: 6-BA, NAA, sucrose, inositol, acid-hydrolyzed casein, and Gynostemma pentaphyllum extract.
[0008] Preferably, the primary induction medium is based on MS medium and contains the following components at mass concentrations: 1.0-2.0 mg / L 6-BA, 0.1-0.5 mg / L NAA, 28-32 g / L sucrose, 100-150 mg / L inositol, 50-80 mg / L acid-hydrolyzed casein, and 0.3-0.6 g / L Gynostemma pentaphyllum extract, with the pH adjusted to 5.8-6.0.
[0009] This invention provides a multi-component synergistic compound system for primary induction culture medium, which can balance the dual needs of bud differentiation and seedling vigor, avoiding problems such as excessive growth or excessive proliferation of callus tissue. At the same time, it specifically adds membrane stabilizing substances, organic nitrogen source supply components and natural antioxidant extracts. The components form a complementary nutrition and regulation network, which can not only ensure nutrient supply and metabolic stability during seed germination, but also induce root primordia development in advance, laying a high-quality seedling foundation for efficient rooting.
[0010] Preferably, the sterilized seeds are inoculated onto a primary induction medium and cultured: During the first week, the temperature is maintained at 24.5-25.0℃, the light intensity at 1200-1300 Lx, the light duration at 10 h / d, and the relative humidity at 70%-75%; during the second week, the temperature is maintained at 24.5-25.0℃, the light intensity is increased to 1300-1500 Lx, the light duration remains at 10 h / d, and the relative humidity is maintained at 70%-75%; during the third week, the temperature is increased to 26.0-26.5℃, the light intensity is increased to 1500-1700 Lx, the light duration is extended to 11 h / d, and the relative humidity is adjusted to 68%-72%; from the fourth week onwards, the temperature is stabilized at 25-25.0℃, the light intensity is increased to 1700-1800 Lx, the light duration is extended to 12 h / d, and the relative humidity is maintained at 68%-72%.
[0011] This invention designs phased cultivation conditions that align with the physiological rhythms of the entire growth cycle of cat's claw grass seeds, from germination to seedling development. Based on the differences in energy requirements and photosynthetic characteristics at different growth stages, a gradient-progressive temperature, light, and humidity control strategy is employed. This condition control mode precisely matches the growth needs of cat's claw grass seeds at different stages, enhancing the plant's adaptability to subsequent cultivation stages.
[0012] Preferably, the subculture induction medium is based on MS medium and contains the following components at mass concentrations: 1.5 mg / L 6-BA, 0.2 mg / L NAA, 30 g / L sucrose and 3.5 g / L agar, with a pH of 5.9.
[0013] Preferably, the proliferation medium is based on MS medium and contains the following components at mass concentrations: 0.5-2.0 mg / L 6-BA, 0.2-1 mg / L NAA, and the pH is adjusted to 5.8-6.0.
[0014] Preferably, the rooting medium is based on 1 / 2 MS medium, with the following components added at mass concentrations: 1-2 mg / L LBA, 0.2-1.0 mg / L NAA, 18-22 g / L sucrose, and the pH is adjusted to 5.8-6.0.
[0015] Preferably, the rooting medium is based on 1 / 2 MS medium, with the following components added at mass concentrations: 0.5 mg / L IBA, 0.2 mg / L NAA, and the pH adjusted to 5.9.
[0016] Preferably, the rooting culture conditions are: a temperature of 23-27℃, a light intensity of 1500-2000 Lx, and a light duration of 12 h / d.
[0017] Preferably, the hardening-off period is 1-2 weeks. Days 1-3: temperature 23-25℃, relative humidity 70%-75%, light 2000-2500Lx, 12h / d, keep the tissue culture bottle sealing film partially open; Days 4-7: temperature 23-25℃, relative humidity 60%-65%, light 2500-3500Lx, 13h / d, remove the sealing film; Days 8-14: temperature 23-25℃, relative humidity 50%-55%, light 3500-5000Lx, 14h / d, natural ventilation.
[0018] Compared with the prior art, the advantages and beneficial effects of the present invention are as follows: 1. This invention provides a multi-component synergistic compound system for primary induction culture medium, which can balance the dual needs of bud differentiation and seedling vigor, avoiding problems such as excessive growth or excessive proliferation of callus tissue. At the same time, it specifically adds membrane stabilizing substances, organic nitrogen source supply components and natural antioxidant extracts. The components form a complementary nutrition and regulation network, which can not only ensure nutrient supply and metabolic stability during seed germination, but also induce root primordia development in advance, laying a high-quality seedling foundation for efficient rooting in the future.
[0019] 2. This invention designs phased cultivation conditions that conform to the physiological rhythm of the entire life cycle from cat's claw seed germination to seedling growth. Based on the differences in energy requirements and photosynthetic characteristics at different growth stages, a gradient progressive temperature, light, and humidity control strategy is adopted. This condition control mode precisely matches the growth needs of cat's claw seeds at different stages, improving the plant's adaptability to subsequent cultivation stages.
[0020] 3. The primary culture medium compound system and the staged culture conditions of this invention have a synergistic effect, producing a synergistic enhancement effect. The precisely compounded culture medium provides basic material support and regulatory signals for plant growth, while the culture conditions adapted to the growth rhythm maximize the functional efficacy of the culture medium components, promote nutrient absorption, hormone signal transduction and antioxidant mechanism activation, accelerate root primordia development and seedling growth, fundamentally solving the core contradiction in the prior art that it is difficult to balance rooting rate and propagation quality. Attached Figure Description
[0021] Figure 1 This is a schematic diagram of the proliferation results in Examples 1-3.
[0022] Figure 2 This is a schematic diagram of the rooting results in Examples 1-10.
[0023] Figure 3 This is a bottom schematic diagram of the rooting results of Examples 1-10.
[0024] Figure 4 This is a schematic diagram of the transplanting results in Examples 1-10. Detailed Implementation
[0025] The technical solutions in the embodiments of the present invention will be clearly and completely described 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.
[0026] All raw materials used in the following embodiments of the present invention are commercially available products: Acid hydrolysis of casein, Gynostemma pentaphyllum extract, Langlin product number: NAT-123, product specification: 90wt% Gynostemma pentaphyllum saponins. Hunan Langlin Bioresources Co., Ltd.
[0027] Yucca extract, Langlin product number: NAT-550, product specification: 60wt% yucca saponins. Hunan Langlin Bioresources Co., Ltd.
[0028] Astragalus extract, Langlin product number: NAT-018, product specification: 50wt% polysaccharide. Hunan Langlin Bioresources Co., Ltd.
[0029] Acid-hydrolyzed casein, CAS: 65072-00-6, Hubei Yuanmeng Biotechnology Co., Ltd.
[0030] Yeast extract, Guangdong Hongyou Biotechnology Co., Ltd.
[0031] For pH adjustment: Acid solution: 0.1 mol / L hydrochloric acid aqueous solution; Alkaline solution: 0.1 mol / L sodium hydroxide aqueous solution.
[0032] Example 1 This embodiment provides a method for rapid propagation of cat's claw grass tissue, including the following steps: (1) Seed selection and disinfection: Select viable seeds, rinse the seeds with running water, soak them in 0.1 (w / v) mercuric chloride (mercuric chloride aqueous solution) on a clean bench for 30 seconds, disinfect them with alcohol for 30 seconds, and finally rinse them with sterile water 4 times.
[0033] (2) Preparation of induction medium: The primary induction medium was based on MS medium, with the following components added at mass concentrations: 1.3 mg / L 6-BA, 0.2 mg / L NAA, 30 g / L sucrose, 120 mg / L inositol, 60 mg / L acid-hydrolyzed casein, and 0.5 g / L Gynostemma pentaphyllum extract, and the pH was adjusted to 5.9. The induction medium was used for seed germination induction; approximately 10 seeds were inoculated per bottle, and 50 bottles were inoculated.
[0034] (3) Seed induction and propagation: The sterilized seeds were inoculated onto the induction medium and cultured. Gradual progressive culture conditions were used to control the conditions. In the first week, the temperature was maintained at 24.5℃, the light intensity at 1200 Lx, the light duration at 10 h / d, and the relative humidity at 70%. In the second week, the temperature was maintained at 24.5℃, the light intensity was increased to 1500 Lx, the light duration was still maintained at 10 h / d, and the relative humidity was maintained at 70%. In the third week, the temperature was increased to 25.5℃, the light intensity was further increased to 1700 Lx, the light duration was extended to 11 h / d, and the relative humidity was adjusted to 72%. By increasing the temperature and light and extending the daylight, the accumulation of photosynthetic products was accelerated, and the seedlings were strengthened. At the same time, the humidity was appropriately reduced to avoid seedling etiolation. In the fourth week and thereafter, the temperature was stabilized at 25.5℃, the light intensity was increased to 1800 Lx, the light duration was extended to 12 h / d, and the relative humidity was maintained at 72%. After 4 weeks of culture, the seeds germinated into seedlings. The seedlings were transferred to fresh subculture induction medium for proliferation culture, and subcultured every 3 weeks (see...). Figure 1 The subculture induction medium was based on MS medium and supplemented with the following components at mass concentrations: 1.5 mg / L 6-BA, 0.2 mg / L NAA, 30 g / L sucrose and 3.5 g / L agar, with a pH of 5.9.
[0035] Screening of proliferation media: 6-Benzylaminopurine (6-BA) and α-naphthaleneacetic acid (NAA) were selected as selection factors. Aseptic seedlings obtained after seed germination were inoculated into proliferation medium, with 6 bottles inoculated for each treatment, and 5 seedlings inoculated per bottle. After 30 days of culture, the number of differentiated shoots in each treatment was measured (differentiation results are shown in Table 2). The proliferation medium was based on MS medium, supplemented with the following components at mass concentrations: 0.5-2.0 mg / L 6-BA, 0.2-1 mg / L NAA, and the pH was adjusted to 5.8-6.0 (design table is shown in Table 1).
[0036] Table 1 Screening of proliferation media
[0037] Table 2 Differentiation results in different proliferation media
[0038] The proliferation medium was based on MS medium, with the following components added at mass concentrations: 1.5 mg / L 6-BA and 0.2 mg / L NAA, and the pH was adjusted to 5.9. This medium was used as the optimal proliferation medium, and step (4) was continued.
[0039] (4) Rooting culture: The propagated seedlings were transferred to a rooting medium based on 1 / 2 MS medium, supplemented with the following components at mass concentrations: 1-2 mg / L IBA, 0.2-1.0 mg / L NAA, and 20 g / L sucrose, and the pH was adjusted to 5.8-6.0. Under the same culture conditions, the adventitious buds could root and form complete plants after 2-3 weeks of culture (see...). Figure 2 , Figure 3 ).
[0040] Screening of rooting culture media: Sucrose, NAA, IBA, and 1 / 2 MS basal medium were selected as selection factors (see Table 3 for the design table). Rooting culture was carried out when the shoot clusters reached a height of 3 cm or more. After 5 days of culture in a medium containing auxin, the rooted seedlings were transferred to the basal medium. After 14 days, the total number of roots and plant growth status were counted (see Table 4).
[0041] Table 3 Screening of rooting culture media
[0042] Table 4 Rooting Results
[0043] The rooting medium was based on 1 / 2 MS medium, with the following components added at different concentrations: 0.5 mg / L IBA and 0.2 mg / L NAA, and the pH was adjusted to 5.9. This medium was the optimal rooting medium, and step (5) was continued.
[0044] (5) Hardening off and transplanting: After rooting, the complete plants were moved to a greenhouse for hardening off. Hardening off for days 1-3: temperature 23℃, relative humidity 70%, light 2000Lx, 12h / d, with the sealing film of the tissue culture bottle partially open; Days 4-7: temperature 23℃, relative humidity 65%, light 2500Lx, 13h / d, with the sealing film removed; Days 8-14: temperature 25℃, relative humidity 55%, light 5000Lx, 14h / d, with natural ventilation. After 2 weeks of hardening off, the plants were transplanted into peat moss, maintaining suitable temperature and humidity, and managed as usual. The growth of the plants was observed after 14 days (see...). Figure 4 ).
[0045] Example 2 The difference between this embodiment and Example 1 is that the primary induction medium is based on MS medium and contains the following components at mass concentrations: 1.0 mg / L 6-BA, 0.5 mg / L NAA, 30 g / L sucrose, 100 mg / L inositol, 80 mg / L acid-hydrolyzed casein, and 0.6 g / L Gynostemma pentaphyllum extract, with the pH adjusted to 5.9.
[0046] Comparative Example 1 The difference between this comparative example and Example 1 is that the Gynostemma pentaphyllum extract was replaced with Yucca extract.
[0047] Comparative Example 2 The difference between this comparative example and Example 1 is that the Gynostemma pentaphyllum extract was replaced with Astragalus membranaceus extract.
[0048] Comparative Example 3 The difference between this comparative example and Example 1 is that the primary induction medium is based on MS medium and contains the following components at mass concentrations: 1.3 mg / L 6-BA, 0.2 mg / L NAA, 30 g / L sucrose, 80 mg / L inositol, 100 mg / L acid-hydrolyzed casein, and 0.8 g / L Gynostemma pentaphyllum extract, with the pH adjusted to 5.9.
[0049] Comparative Example 4 The difference between this comparative example and Example 1 is that the primary induction medium is based on MS medium and contains the following components at mass concentrations: 1.3 mg / L 6-BA, 0.2 mg / L NAA, 30 g / L sucrose, 170 mg / L inositol, 30 mg / L acid-hydrolyzed casein, and 0.1 g / L Gynostemma pentaphyllum extract, with the pH adjusted to 5.9.
[0050] Comparative Example 5 The difference between this comparative example and Example 1 is that the primary induction medium is based on MS medium and contains the following components at mass concentrations: 1.3 mg / L 6-BA, 0.2 mg / L NAA, 30 g / L sucrose, 170 mg / L mannitol, 0.2 g / L yeast extract, and 0.1 g / L Gynostemma pentaphyllum extract, with the pH adjusted to 5.9.
[0051] Comparative Example 6 The difference between this comparative example and Example 1 is that the sterilized seeds were inoculated onto the primary induction medium and cultured under the conditions of a temperature of 23-27℃, a light intensity of 1500-2000 Lx, and a light duration of 12 h / d.
[0052] Performance testing Following the propagation methods of Examples 1-2 and Comparative Examples 1-6, 10 seeds were inoculated per bottle for each example and comparative example, with 50 bottles inoculated. The rooting rate was calculated, and the average root length after rooting culture was also calculated. Average root length (cm) = total root length (cm) ÷ number of adventitious roots (strips), where total root length (cm) = Σ (length of a single adventitious root). Rooting rate = number of rooted plants ÷ total number of seeds × 100%.
[0053] The results are shown in Table 5.
[0054] Table 5 Performance Test Results
[0055] As shown in Table 1, Examples 1-2 have high rooting rates and long average root lengths.
[0056] In Comparative Example 1, replacing Gynostemma pentaphyllum extract with Yucca extract resulted in a decreased effect. Analysis revealed that the core component of Yucca extract is steroidal saponins, which can improve culture medium permeability and provide broad-spectrum antibacterial activity, but its antioxidant activity and cell division-promoting function are far weaker than those of Gynostemma pentaphyllum extract. In the initial culture of *Cephalotaxus fortunei*, browning is a key factor restricting seedling vigor; Yucca extract cannot effectively alleviate oxidative stress, leading to poor stress resistance and hindered root primordia differentiation in the initial seedlings.
[0057] In Comparative Example 2, replacing Gynostemma pentaphyllum extract with Astragalus membranaceus extract resulted in a decreased efficacy. Analysis revealed that the main active ingredient in Astragalus membranaceus extract is Astragalus polysaccharide, whose core function is to enhance plant stress resistance. However, it lacks the saponins found in Gynostemma pentaphyllum extract, which promote cell division and synergistically function with hormones 6-BA / NAA.
[0058] In Comparative Examples 3-4, the concentration of components was unbalanced, the synergistic effect of the three components decreased, resulting in decreased cell membrane stability, reduced nutrient absorption efficiency, and slow root development. In Comparative Example 5, replacing inositol with mannitol and acid-hydrolyzed casein with yeast extract resulted in a decreased efficacy. Analysis suggests that mannitol can only maintain osmotic pressure and cell membrane stability, but lacks the functions of inositol in regulating carbohydrate metabolism and synergistically modulating hormone signaling, leading to insufficient nutrient accumulation in seedlings. Replacing acid-hydrolyzed casein with yeast extract: While yeast extract contains B vitamins and nucleic acid derivatives, its proportion of small-molecule amino acids is lower than that of acid-hydrolyzed casein, resulting in decreased efficiency in organic nitrogen source supply.
[0059] In Comparative Example 6, the cat's claw grass showed poor adaptability to the physiological needs of plants grown under uniform culture conditions.
[0060] The above description represents the preferred embodiments of the present invention. It should be noted that those skilled in the art can make various improvements and modifications without departing from the principles of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.
Claims
1. A method for rapid propagation of cat's claw plant tissue, characterized in that, Includes the following steps: (1) Seed selection and disinfection: Select viable seeds, wash the seeds, soak them in mercuric chloride, disinfect them with alcohol, and finally rinse them with sterile water. (2) Preparation of primary induction medium: Preparation of primary induction medium; (3) Seed induction and proliferation: The sterilized seeds were inoculated on the primary induction medium and cultured; after 4-6 weeks of culture, the seeds germinated into seedlings; the seedlings were transferred to the subculture induction medium for proliferation culture; (4) Rooting culture: Transfer the propagated seedlings to the rooting culture medium for cultivation; (5) Hardening off and transplanting: After the roots have developed, the complete plants are moved to the greenhouse for hardening off. After hardening off for 1-2 weeks, the plants are transplanted into peat soil.
2. The method for rapid propagation of cat's claw grass tissue according to claim 1, characterized in that, The primary induction medium was based on MS medium and contained the following components: 6-BA, NAA, sucrose, inositol, acid-hydrolyzed casein, and Gynostemma pentaphyllum extract.
3. The method for rapid propagation of cat's claw grass tissue according to claim 2, characterized in that, The primary induction medium was based on MS medium and contained the following components at mass concentrations: 1.0-2.0 mg / L 6-BA, 0.1-0.5 mg / L NAA, 28-32 g / L sucrose, 100-150 mg / L inositol, 50-80 mg / L acid-hydrolyzed casein, and 0.3-0.6 g / L Gynostemma pentaphyllum extract, with the pH adjusted to 5.8-6.
0.
4. The method for rapid propagation of cat's claw grass tissue according to claim 1, characterized in that, The sterilized seeds were inoculated onto the primary induction medium and cultured: During the first week, the temperature was maintained at 24.5-25.0℃, light intensity at 1200-1300 Lx, light duration at 10 h / d, and relative humidity at 70%-75%; during the second week, the temperature was maintained at 24.5-25.0℃, the light intensity was increased to 1300-1500 Lx, the light duration remained at 10 h / d, and the relative humidity was maintained at 70%-75%; during the third week, the temperature was increased to 26.0-26.5℃, the light intensity was increased to 1500-1700 Lx, the light duration was extended to 11 h / d, and the relative humidity was adjusted to 68%-72%. In the fourth week and thereafter, the temperature will be stabilized at 25-25.05℃, the light intensity will be increased to 1700-1800 Lx, the light duration will be extended to 12h / d, and the relative humidity will be maintained at 68%-72%.
5. The method for rapid propagation of cat's claw grass tissue according to claim 1, characterized in that, The subculture induction medium was based on MS medium and supplemented with the following components at mass concentrations: 1.5 mg / L 6-BA, 0.2 mg / L NAA, 30 g / L sucrose and 3.5 g / L agar, with a pH of 5.
9.
6. The method for rapid propagation of cat's claw grass tissue according to claim 1, characterized in that, The proliferation medium is based on MS medium and contains the following components at mass concentrations: 0.5-2.0 mg / L 6-BA and 0.2-1 mg / L NAA, with the pH adjusted to 5.8-6.
0.
7. The method for rapid propagation of cat's claw grass tissue according to claim 1, characterized in that, The rooting medium was based on 1 / 2 MS medium, with the following components added at mass concentrations: 1-2 mg / L IBA, 0.2-1.0 mg / L NAA, 18-22 g / L sucrose, and the pH was adjusted to 5.8-6.
0.
8. The method for rapid propagation of cat's claw grass tissue according to claim 7, characterized in that, The rooting medium was based on 1 / 2 MS medium, with the following components added at different concentrations: 0.5 mg / L IBA and 0.2 mg / L NAA, and the pH was adjusted to 5.
9.
9. The method for rapid propagation of cat's claw grass tissue according to claim 8, characterized in that, The conditions for rooting culture are: temperature of 23-27℃, light intensity of 1500-2000 Lx, and light duration of 12 h / d.
10. The method for rapid propagation of cat's claw grass tissue according to claim 1, characterized in that, The hardening-off period is 1-2 weeks. Days 1-3: Temperature 23-25℃, relative humidity 70%-75%, light 2000-2500Lx, 12h / d, keep the tissue culture bottle sealing film partially open; Days 4-7: Temperature 23-25℃, relative humidity 60%-65%, light 2500-3500Lx, 13h / d, remove the sealing film; Days 8-14: Temperature 23-25℃, relative humidity 50%-55%, light 3500-5000Lx, 14h / d, natural ventilation.