Method for breaking dormancy period of paris polyphylla seeds

Abstract

The invention discloses a method for breaking the dormancy period of paris polyphylla seeds. The method comprises the steps of S1, seed pretreatment, wherein the paris polyphylla seeds are washed and disinfected; s2, soaking treatment: soaking the pretreated paris polyphylla seeds in a gibberellin solution in a dark environment at 22 DEG C; s3, germination accelerating treatment: placing the soaked seeds in a sterile culture dish, alternately placing the seeds in a dark low-temperature environment, and repeatedly accelerating germination of the seeds; S4, rapid propagation: transferring the germinated paris polyphylla seeds subjected to germination accelerating to a basic culture medium, adding NAA and 6BA into the basic culture medium, and meanwhile, cooperating with a light environment, so as to obtain sterile plants. According to the method for breaking the dormancy period of the paris polyphylla seeds, the germination rate is increased, the germination rate of the paris polyphylla seeds is remarkably increased through a gibberellin treatment and cold treatment combined method, and the technical problem that the germination rate of the paris polyphylla seeds is low is solved; the planting period is shortened; the rapid propagation of the paris polyphylla is realized.

Description

Technical Field

[0001] The invention relates to the technical field of Chinese medicinal material planting, in particular to a method for breaking the dormancy period of Paris polyphylla seeds. Background Art

[0002] As an important traditional Chinese medicine, Paris polyphylla var. yunnanensis has a significant dormancy characteristic in its seeds, which severely limits the efficiency and yield of its artificial cultivation. Its dormancy characteristic is mainly manifested in the following aspects:

[0003] Seed coat barrier: The seed coat of Polygonum multiflorum seeds is thick and contains substances that inhibit germination, which hinders the seeds' absorption of water and oxygen, thereby prolonging the dormancy period.

[0004] Embryo after-ripening: The embryo of the seeds of Paris polyphylla is morphologically fully developed, but physiologically it is not yet mature and needs to go through a period of after-ripening before it can germinate.

[0005] Environmental condition restrictions: The seeds of Polygonum multiflorum have high requirements for environmental conditions such as temperature and humidity. It is difficult to break dormancy in an unsuitable environment. The dormancy period of Polygonum multiflorum seeds is relatively long. Under conventional breeding conditions, it takes 8 years of planting to generate economic benefits. The germination rate of untreated Polygonum multiflorum seeds is low, the cost is high, and the sowing is complicated.

[0006] In the existing technology, the seeds of Paris polyphylla usually need to go through a long natural dormancy period (up to several years), and the germination rate of untreated seeds is extremely low, resulting in high planting costs and long cycles, making it difficult to meet market demand. Summary of the Invention

[0007] In view of the deficiencies in the prior art, the present invention provides a method for breaking the dormancy period of Paris polyphylla seeds, aiming to solve the above problems.

[0008] To achieve the above object, the present invention provides the following technical solutions:

[0009] A method for breaking the dormancy period of Paris polyphylla seeds, comprising:

[0010] S1: Seed pretreatment: Wash and disinfect the Paris polyphylla seeds;

[0011] S2: Soaking treatment: The pretreated Paris polyphylla seeds were soaked in a gibberellin solution in a dark environment at 22°C.

[0012] S3: Germination treatment: The soaked seeds are placed in a sterile culture dish and placed alternately in a dark and low temperature environment to repeatedly germinate the seeds.

[0013] S4: Rapid propagation: Transfer the germinated Paris polyphylla seeds to a basic culture medium, add NAA and 6BA to the basic culture medium, and adjust the light environment to obtain sterile plants.

[0014] Preferably, the S1 further includes:

[0015] Step 1: Rinse: Rinse the seeds of Paris polyphylla thoroughly with running water to remove impurities and microorganisms on the surface of the seeds;

[0016] Step 2: Preliminary disinfection: Soak in ethanol to disinfect and kill most bacteria and fungi on the surface of the seeds;

[0017] Step 3: Deep disinfection: After preliminary disinfection, the seeds are soaked in HgCl2 to further kill pathogens on the surface of the seeds.

[0018] Preferably, the ethanol concentration in step 2 is 70%, and the soaking time is 2-5 minutes;

[0019] The HgCl2 concentration in step 2 is 0.2%, and the soaking time is 10-30 minutes.

[0020] Preferably, the concentration of the gibberellin solution in S2 is 20-80 ppm, and the soaking time is 48 hours.

[0021] Preferably, the low temperature environment in S3 is 4° C. and 22° C., the number of alternating placements is 3 times, the placement time is one week, and the number of repetitions is 3-5 times.

[0022] Preferably, the basic culture medium in S4 is a hormone-free 1 / 2MS culture medium, the concentration of NAA is 0.5 mg / L, and the concentration of 6BA is 0.1 mg / L;

[0023] The illumination in S4 lasts for 10 hours per day, and the light intensity is 1000 Lux.

[0024] Compared with the prior art, the present invention has the following beneficial effects:

[0025] The present invention provides a method for breaking the dormancy period of Paris polyphylla seeds, and the technical effects are summarized as follows:

[0026] Improve germination rate: By combining gibberellin treatment and cold treatment, the germination rate of Paris polyphylla seeds was significantly improved, up to 100%. This solved the technical problem of low germination rate of Paris polyphylla seeds, provided a reliable source of seeds for artificial planting, ensured the supply of Paris polyphylla medicinal materials, and overcame the shortcomings of Paris polyphylla medicinal materials in existing technologies, such as poor germination and slow proliferation of sexual seedlings.

[0027] Shorten the planting cycle: The rapid reproduction of Polygonum multiflorum is achieved, the planting cycle is shortened, and the planting efficiency is improved. By optimizing the technical solutions, the Polygonum multiflorum plants can reach the commercialization standards in a shorter time, thus meeting the market demand.

[0028] Reduce costs: It improves the germination rate and reproduction speed, reduces the planting cost and management difficulty. By reducing the amount of seeds used, shortening the planting cycle and reducing the management difficulty, the artificial planting cost of Polygonum multiflorum can be significantly reduced.

[0029] Promote genetic breeding: It provides effective technical support for the genetic breeding of Paris polyphylla, which is conducive to the cultivation of high-yield, high-quality new Paris polyphylla varieties. By applying this technical solution, the genetic improvement process of Paris polyphylla can be accelerated, and new varieties with greater market competitiveness can be cultivated. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 Schematic diagram of the process of the present invention. DETAILED DESCRIPTION

[0031] The following will clearly and completely describe the technical solutions in the embodiments of the present invention in conjunction with the accompanying drawings. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative efforts are within the scope of protection of the present invention.

[0032] The embodiment proposed in this application is: a method for breaking the dormancy period of Paris polyphylla seeds, comprising the following method:

[0033] S1: Seed pretreatment:

[0034] Step 1: Rinse: Rinse the seeds thoroughly with running water to remove impurities and microorganisms on the surface. This step is crucial to reduce pathogens and impurities on the seed surface and helps improve the germination rate of the seeds.

[0035] Step 2: Preliminary disinfection: Soak in 70% ethanol for 2-5 minutes for preliminary disinfection. Ethanol can effectively kill most bacteria and fungi on the surface of the seeds, laying the foundation for subsequent deep disinfection.

[0036] Step 3: Deep disinfection: Soak the seeds that have undergone preliminary disinfection in 0.2% HgCl2 (mercuric chloride) for 10-30 minutes to further kill the pathogens on the surface of the seeds. Mercuric chloride, as a powerful disinfectant, can completely kill various pathogens on the surface of the seeds to ensure the sterility of the seeds.

[0037] S2: Soaking treatment:

[0038] The pre-treated and disinfected seeds are placed in a 20-80ppm concentration of gibberellin solution for soaking in the dark at 22°C for 48 hours. Gibberellic acid, as a plant growth regulator, can promote seed germination and growth. Treatment with an appropriate concentration of gibberellin can significantly improve the subsequent germination rate of Dendrobium officinale seeds.

[0039] S3: Germination treatment:

[0040] The soaked seeds are placed in a sterile culture dish and placed alternately at 4°C and 22°C in the dark for 1 week, and repeated 3-5 times for seed germination. Cold treatment can simulate the low temperature conditions in the natural environment, which helps to break the dormancy of the seeds, promote seed germination, and germinate.

[0041] S4: Rapid reproduction:

[0042] The germinated seeds of Paris polyphylla after germination were transferred to a hormone-free 1 / 2MS medium, and the seeds were cultivated with 1 / 2MS as the basic medium. 0.5 mg / L of NAA (naphthaleneacetic acid) and 0.1 mg / L of 6BA (6-benzylaminopurine) were added to the basic medium for rapid propagation. The cultivation process was coordinated with the light environment. Sterile plants were obtained by culturing under light. The light lasted for 10 hours a day with a light intensity of 1000 Lux. 1 / 2MS medium, as a commonly used plant tissue culture medium, can provide various nutrients required for plant growth. Light is one of the important factors for plant growth. Suitable light conditions can promote photosynthesis and metabolic activities of seeds, thereby further accelerating seed germination and growth. By adding appropriate concentrations of NAA and 6BA, the rapid propagation and growth of Paris polyphylla seeds can be promoted. NAA and 6BA are commonly used plant growth regulators. By adjusting their concentration ratio, the growth conditions of Paris polyphylla plants can be optimized, and the proliferation of buds and the growth of roots can be promoted.

[0043] Experimental data and analysis:

[0044] Effects of cold treatment and light on germination rate:

[0045] Three discontinuous cold treatments (4°C and 22°C alternating) significantly increased the germination rate of seeds;

[0046] The germination rate of continuous 22℃ room temperature treatment is low. Although the seeds can germinate, the germination rate is lower than that of cold treatment.

[0047] Continuous low temperature treatment at 4°C had no significant effect and could not increase the germination rate of seeds;

[0048] Light treatment at room temperature is not conducive to seed germination;

[0049] The above contents are shown in Table 1, which indicates that cold treatment and dark conditions are essential for breaking the dormancy of R. yunnanensis seeds.

[0050] Treatment method Germination rate after 60 days (%) 120-day statistical germination rate (%) 3 discontinuous dark cooling treatments 60 90 Continuously treat in dark place at room temperature of 22℃ 30 50 Continuous 4℃ low temperature dark treatment 10 20 Light treatment at room temperature 20 30

[0051] Table 1

[0052] Effect of gibberellin concentration on germination rate:

[0053] The experimental results showed that, as shown in Table 2, the appropriate concentration of gibberellin (20-80 ppm) treatment can significantly increase the germination rate of Paris polyphylla seeds, the highest of which can approach 100%, which indicates that gibberellin treatment is one of the effective methods to break the dormancy of Paris polyphylla seeds;

[0054] The experiment on the effect of gibberellins on the germination of Polygonum multiflorum seeds was carried out on the basis that cold treatment has a significant promoting effect on seed germination. It can be seen from Table 2 that appropriate gibberellins (20-80ppm) treatment and germination in the dark at room temperature (22℃) can achieve similar results to cold treatment. If appropriate concentration of gibberellins is used for soaking and discontinuous cold treatment is performed on the seeds at the same time, the germination rate of the seeds can be more effectively improved.

[0055] From Table 1 and Table 2, we can conclude that the seeds of Paris polyphylla germinated continuously for 60 days and 120 days at room temperature (22°C) and 4°C, and the germination rate was below 50%, indicating that the seeds were dormant. Both cold treatment and gibberellin treatment could break the dormancy of Paris polyphylla seeds.

[0056] And light is not conducive to seed germination.

[0057]

[0058] Table 2

[0059] Effects of different culture media on micropropagation:

[0060] As shown in Table 3, the 1 / 2MS medium supplemented with 0.5 mg / L NAA and 0.1 mg / L 6BA was beneficial to both bud proliferation and root growth, indicating that this medium formula was a suitable choice for the rapid propagation of Paris polyphylla. Preliminary experimental results showed that reducing the inorganic salt content was beneficial to the growth of sterile seedlings.

[0061] culture medium Observation results 1 / 2MS+NAAO.5og / L The plant grows roots but does not proliferate buds. 1 / 2MS+NAAO.5ng / L+6BA0.1ng / L The plant is relatively small, with many buds and growing roots. 1 / 2MS+NAA0.25ag / L+6BA1.0mg / L The plant is sturdy, the rhizome is enlarged, and the bud proliferation is small 1 / 2MS+6BA1.0mg / L Root lotus swelling, no bud proliferation

[0062] Table 3

[0063] In another embodiment, NAA and 6BA in S4 are replaced by oligosaccharides PP-DP7, PP-DP8 and PP-DP9 to study their effects on the proliferation of Paris polyphylla buds. Oligosaccharides, as a new type of plant growth regulator, have the function of promoting plant growth and development. By studying the effects of different concentrations of oligosaccharides on the proliferation of Paris polyphylla buds, more effective bud proliferation promoters can be found.

[0064] Effects of oligosaccharides on bud proliferation:

[0065] As shown in Table 4, oligosaccharides have no obvious effect on root differentiation and growth, but can affect bud proliferation. The optimal concentrations of PP-DP7, PP-DP8 and PP-DP9 that are beneficial to bud proliferation are 2.5ppm, 10ppm and 20ppm, respectively. This shows that different concentrations of oligosaccharides have different promoting effects on the proliferation of Paris polyphylla buds, and the appropriate type and concentration of oligosaccharides can be selected according to actual needs.

[0066]

[0067] Table 4

[0068] Through the above method, the rapid propagation of Paris polyphylla can be achieved, and the annual growth of Paris polyphylla rhizomes can be increased by using biotechnology, so as to facilitate the planned development of Paris polyphylla planting industry, ensure the supply of Paris polyphylla medicinal materials, and overcome the passive situation of Paris polyphylla medicinal materials relying entirely on wild resources.

[0069] Through temperature regulation and gibberellin regulation, the dormancy period of the seeds of Paris polyphylla can be effectively broken, the seed germination rate can be improved, and the shortcomings of the existing technology of poor germination and slow proliferation of sexual seedlings can be overcome.

[0070] Compared with the existing technology, it has the advantages of fast reproduction speed and short cycle, so as to realize industrial rapid reproduction.

[0071] It is conducive to further research and industrialization of the genetic breeding of Paris polyphylla (such as the establishment of strains with high steroid saponin content, high polysaccharide and oligosaccharide content) and physiological and biochemical (such as the relationship between gelatin and powdery Paris polyphylla, the relationship between microbial community and Paris polyphylla quality, etc.).

[0072] It should be noted that, in this document, relational terms such as first and second, etc., are used only to distinguish one entity or operation from another entity or operation, and do not necessarily require or imply the existence of any such actual relationship or order between these entities or operations. Moreover, the terms "comprises," "comprising," or any other variants thereof are intended to cover non-exclusive inclusion, so that a process, method, article, or device comprising a series of elements includes not only those elements, but also other elements not explicitly listed, or elements inherent to such process, method, article, or device. In the absence of further limitations, an element defined by the phrase "comprising a ..." does not exclude the presence of other identical elements in the process, method, article, or device comprising the element.

[0073] While embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions, and variations may be made to these embodiments without departing from the principles and spirit of the invention, and that the scope of the invention is defined by the appended claims and their equivalents.

Claims

1. A method for breaking the dormancy period of Paris polyphylla seeds, characterized in that: include: S1: Seed pretreatment: Wash and disinfect the Paris polyphylla seeds; S2: Soaking treatment: The pretreated Paris polyphylla seeds were soaked in a gibberellin solution in a dark environment at 22°C. S3: Germination treatment: The soaked seeds are placed in a sterile culture dish and placed alternately in a dark and low temperature environment to repeatedly germinate the seeds. S4: Rapid propagation: Transfer the germinated Paris polyphylla seeds to a basic culture medium, add NAA and 6BA to the basic culture medium, and adjust the light environment to obtain sterile plants.

2. A method for breaking the dormancy period of Paris polyphylla seeds according to claim 1, characterized in that: Said S1 further comprises: Step 1: Rinse: Rinse the seeds of Paris polyphylla thoroughly with running water to remove impurities and microorganisms on the surface of the seeds; Step 2: Preliminary disinfection: Soak in ethanol to disinfect and kill most bacteria and fungi on the surface of the seeds; Step 3: Deep disinfection: After preliminary disinfection, the seeds are soaked in HgCl2 to further kill pathogens on the surface of the seeds.

3. A method for breaking the dormancy period of Paris polyphylla seeds according to claim 2, characterized in that: The ethanol concentration in step 2 is 70%, and the soaking time is 2-5 minutes; The HgCl2 concentration in step 2 is 0.2%, and the soaking time is 10-30 minutes.

4. A method for breaking the dormancy period of Paris polyphylla seeds according to claim 1, characterized in that: The concentration of the gibberellin solution in S2 is 20-80 ppm, and the soaking time is 48 hours.

5. The method for breaking the dormancy period of Paris polyphylla seeds according to claim 1, wherein: The low temperature environment in S3 is 4° C. and 22° C., the number of alternating placements is 3 times, the placement time is one week, and the number of repetitions is 3-5 times.

6. The method for breaking the dormancy period of Paris polyphylla seeds according to claim 1, wherein: The basic medium in the S4 is a hormone-free 1 / 2MS medium, the concentration of NAA is 0.5 mg / L, and the concentration of 6BA is 0.1 mg / L; The illumination in S4 lasts for 10 hours per day, and the light intensity is 1000 Lux.

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