A method for improving the survival rate of paeony test-tube seedling transplanting

Abstract

The application discloses a method for improving the survival rate of Paeonia suffruticosa test tube seedlings, which comprises the following steps: mixing grass carbon, perlite and vermiculite to prepare a mixed substrate, transplanting the Paeonia suffruticosa test tube seedlings into a nutrient pot filled with the mixed substrate, and performing post-planting management, wherein the relative water content of the mixed substrate is kept at 56-75% during the post-planting management, and the environmental temperature is controlled at 12-18 DEG C. The application determines the temperature and the water content of the substrate which affect the survival of the Paeonia suffruticosa test tube seedlings, and the survival rate of the Paeonia suffruticosa test tube seedlings is increased from 35% to 60%. By adding a self-developed microbial preparation, the hardening time is shortened under the suitable temperature and the water content of the substrate, and strong seedlings are obtained, which not only solves the rot problem of the rhizome part of the Paeonia suffruticosa test tube seedlings during the transplanting, but also promotes the rapid growth of the leaves and petioles of the above-ground part, and the survival rate of the Paeonia suffruticosa test tube seedlings is improved to 80%, thereby laying a foundation for realizing the large-scale transplanting of the Paeonia suffruticosa test tube seedlings.

Description

Technical Field

[0001] This invention relates to the field of peony cultivation, and in particular to a method for improving the survival rate of peony seedlings transplanted from test tubes. Background Technology

[0002] Peony ( Paeonia lactiflora Peony (Paeonia lactiflora) is a perennial herbaceous plant belonging to the genus Paeonia in the family Paeoniaceae. It is a traditional famous flower in my country. Traditional peony cultivation techniques mainly rely on sowing and division for propagation, resulting in a low propagation coefficient that makes it difficult to meet market demand and hinders the industrialization of peony cultivation.

[0003] Plant tissue culture technology utilizes isolated plant organs (such as roots and stems), tissues (such as epidermis and endosperm), or cells (such as megaspores, microspores, and somatic cells), as well as protoplasts, under sterile conditions and suitable artificial culture media and temperatures. This process induces callus formation, adventitious buds, and adventitious roots, ultimately leading to complete plants. Plant tissue culture technology offers advantages such as high efficiency, rapid production, and year-round production, making it an important method for promoting the production of superior varieties and the development of the plant industry.

[0004] There are many research reports on peony tissue culture technology. Currently, the proliferation culture of peony test-tube seedlings has been achieved. There are a few reports on the rooting of test-tube seedlings, mostly focusing on the effects of auxin type and concentration on rooting. However, the transplanting of rooted peony test-tube seedlings has not been successful. For example, the "Induction and Rooting Technology of Adventitious Buds in Ornamental Peony 'Da Fu Gui'" (Wu Hongjuan et al., Journal of Northeast Forestry University, 2011, 39(9):20–22) was propagated by inducing adventitious bud formation, but browning and vitrification occurred, with a vitrification rate as high as 40.91%. In addition, the transplanting and cultivation of rooted peony test-tube seedlings was also disclosed. After transplanting, the seedlings were managed in an environment of 22℃ and 80% humidity, and nutrient solution was sprayed once a week. The results showed that the rootstock of the peony rotted severely after transplanting and the above-ground parts stopped growing. Only a few relatively strong tissue culture seedlings survived for 3 weeks after transplanting, but no new leaves grew, and they gradually died afterward. It can be seen that this report did not achieve the transplanting of peony test-tube seedlings.

[0005] Chinese patent ZL 202110624827.X discloses a method for propagating peonies using tissue culture technology (this method is an early application of the applicant). This method uses a test-tube dormant bud approach for rapid propagation of peonies through tissue culture, achieving a high bud disc structure induction rate (above 90%) and minimal vitrification, thus improving the rooting rate and quality of test-tube rooted seedlings and obtaining robust rooted seedlings. However, this patent does not achieve the transplantation of peony test-tube seedlings.

[0006] In summary, the root rot and cessation of above-ground growth of peony in vitro seedlings after transplanting remain significant challenges. Therefore, resolving the root rot problem and ensuring normal above-ground growth of peony seedlings after transplanting is crucial for the large-scale cultivation of peony in vitro seedlings. Summary of the Invention

[0007] In view of this, the purpose of this invention is to provide a method to improve the survival rate of peony test-tube seedlings after transplanting, which solves the technical problem of peony test-tube seedlings failing to survive after transplanting and improves the peony tissue culture rapid propagation technology system.

[0008] To achieve the above objectives, the present invention adopts the following technical solution:

[0009] The method for improving the survival rate of peony test-tube seedlings after transplanting according to the present invention is as follows: using a mixed substrate made of peat moss, perlite and vermiculite as the transplanting substrate, transplanting the peony test-tube seedlings into nutrient pots containing the transplanting substrate, and performing post-transplanting management. During the post-transplanting management process, the moisture content of the transplanting substrate is maintained at 56% to 75%, and the post-transplanting ambient temperature is controlled at 12 to 18°C.

[0010] In the above-mentioned solution, the present invention addresses the defects of peony test-tube seedlings, such as easy rotting of the rhizome and cessation of growth of the above-ground parts after transplanting. By rationally selecting the growth conditions after transplanting, the invention promotes the rapid recovery of the root system of peony test-tube seedlings, shortens the recovery time, improves the transplant survival rate of peony test-tube seedlings, solves the technical problem of peony test-tube seedlings failing to survive after transplanting, and improves the rapid propagation technology system of peony tissue culture.

[0011] In this invention, the volume ratio of peat moss, perlite and vermiculite in the transplanting substrate is 2:2:1.

[0012] In this invention, the transplanting substrate further includes water or a microbial preparation. The preparation steps of the transplanting substrate include: mixing peat moss, perlite, and vermiculite in a volume ratio of 2:2:1, and then mixing with water or a microbial preparation. The microbial preparation is a Bacillus subtilis inoculant or a microbial growth solution. The effective viable count of Bacillus subtilis in the Bacillus subtilis inoculant is 0.8–1.2 billion / mL; the effective viable count of Bacillus subtilis in the microbial growth solution is ≥2.0 billion / mL. More preferably, the effective bacteria in the Bacillus subtilis inoculant are Bacillus subtilis Pro1A2.

[0013] In nature, the plant growth environment contains a large number of microorganisms. These microorganisms can break down organic matter in the growth substrate into humus that can be absorbed by plants, promoting root and above-ground growth. Microorganisms can also secrete chitinase, preventing the invasion of harmful bacteria and thus preventing pests and diseases. Therefore, in hydroponics, microbial strains can be artificially added to promote plant growth. While there are reports of microorganisms promoting plant growth, there is currently no research on peony. This invention, through the rational selection of post-planting temperature and substrate moisture content combined with microbial agents, significantly shortens the seedling establishment time and cultivates robust seedlings. It not only solves the problem of root and stem rot in peony test-tube seedlings during transplanting but also promotes rapid growth of leaves and petioles, increasing the survival rate of transplanted peony test-tube seedlings (up to 80%). This lays the foundation for large-scale transplanting of peony test-tube seedlings and has significant promotional value.

[0014] In this invention, during the post-planting management process, the plant is initially kept in a sealed, moist environment, followed by ventilation management 12–18 days after planting. More preferably, ventilation begins gradually 15 days after the peony test-tube seedlings are transplanted.

[0015] In this invention, the transplanting temperature during the post-planting management process is preferably controlled at 15℃ to ensure that the root system of peony test-tube seedlings recovers and grows rapidly, reduce the reproduction rate of pathogens, and improve the transplant survival rate of peony test-tube seedlings.

[0016] Furthermore, the moisture content of the transplanting substrate during post-planting management is preferably 61%–75%. More preferably, the moisture content of the transplanting substrate is consistently controlled at 66%–70% during post-planting management. During plant growth, the moisture content of the substrate is a crucial factor in ensuring normal plant growth. This invention not only prevents root rot and death of peony seedlings due to overly wet substrate, but also prevents browning of the root tips of peony seedlings due to overly dry transplanting substrate.

[0017] The innovation of this invention lies in the following: Addressing the issue of easy rotting and death of peony test-tube seedlings after transplanting, this invention solves the problem of seedling rotting and death by regulating environmental factors (temperature and substrate moisture content); this invention clarifies the temperature and substrate moisture content parameters affecting the survival rate of peony test-tube seedlings after transplanting, increasing the survival rate from 35% to 60%; by adding a self-developed microbial agent, under suitable temperature and substrate moisture conditions, the seedling recovery time is shortened, resulting in robust seedlings. This not only solves the problem of root and stem rotting during transplanting but also promotes the rapid growth of leaves and petioles above ground, increasing the survival rate of peony test-tube seedlings to 80%, laying the foundation for large-scale transplanting of peony test-tube seedlings. Attached Figure Description

[0018] Figure 1This describes the growth of experimental group 2 in Example 1 of this invention during post-planting management.

[0019] Figure 2 This describes the root growth of control group 1 and experimental group 2 after transplanting in Example 1 of this invention.

[0020] Figure 3 This describes the growth of peony plants in three treatment groups in Example 3 of this invention after 60 days of post-planting management. Detailed Implementation

[0021] The method of the present invention will be described in more detail below through specific embodiments to facilitate understanding by those skilled in the art. It should be noted that, unless otherwise specified, the reagents used in the following embodiments are commonly used laboratory reagents, and the methods employed are conventional methods in the art.

[0022] The method for improving the survival rate of peony test-tube seedlings according to the present invention includes the following steps: transplanting peony test-tube seedlings using a mixed substrate of peat moss, perlite, and vermiculite as the transplanting substrate; transplanting the seedlings into nutrient pots containing the transplanting substrate; and placing the nutrient pots in a plant cultivation box for post-planting management. During post-planting management, the moisture content of the transplanting substrate is maintained at 56%–75%, and the ambient temperature after transplanting is controlled at 12–18℃. It should be noted that the moisture content of the transplanting substrate in this invention is determined using a conventional drying method.

[0023] Example 1: Effect of post-planting temperature on the survival rate of peony test-tube seedlings after transplanting

[0024] The first step is to mix peat moss, perlite, and vermiculite in a volume ratio of 2:2:1, and then mix them evenly with tap water to obtain a transplanting substrate with a moisture content of 61-65%.

[0025] The second step is to fill the transplanting substrate into the nutrient pots, transplant the peony test-tube seedlings into each nutrient pot, and place the nutrient pots in different plant culture boxes. The transplanted peony test-tube seedlings are divided into four groups, including experimental group 1, experimental group 2, experimental group 3 and control group 1.

[0026] The third step is to manage the transplanted peony seedlings after transplanting. In the early stage after transplanting, the four groups of peonies are kept moist. Fifteen days after transplanting, water them appropriately during the post-transplanting management process to keep the moisture of the transplanting substrate at 61-65%.

[0027] In addition, the plant cultivation box was kept at a constant temperature during post-planting management. The post-planting management temperatures for experimental groups 1, 2, and 3 were 12℃, 15℃, and 18℃, respectively; the post-planting management temperature for control group 1 was the conventional management temperature for existing seedling transplanting (i.e., 25℃).

[0028] The rooting and survival rates of each group of plants were statistically analyzed using standard methods. The survival rates of experimental groups 1-3 and control group 1 60 days after transplanting are shown in Table 1. The growth status of experimental group 2 is shown in [Table 1]. Figure 1 The root system conditions of control group 1 and experimental group 2 are shown in the figure. Figure 2 .

[0029] Table 1. Survival rate of four groups of peony plants in Example 1 of the present invention.

[0030]

[0031] The growth of the above-mentioned peony plants during post-planting management is as follows: (Combined with...) Figure 1-2 It can be seen that the leaves of experimental groups 1-3 grew relatively slowly, with the leaves turning green first. The plants in these groups began to grow 15 days after transplanting, and by day 15, some plants in the experimental groups had already developed root systems. After 30 days of transplanting, the leaves were fully expanded and the root systems were growing well. The root and leaf growth conditions are shown in the table below. Figure 2 The leaves grew significantly around 45 days after transplanting; and some peony plants survived and developed well-developed root systems 60 days after transplanting, as shown in Table 1. The results indicate that this invention not only reduces the rate of pathogen reproduction but also ensures that the roots of the peony plantlets grow before the leaves after transplanting, thus improving the survival rate of the transplanted peony plantlets and laying the foundation for large-scale transplanting of peony plantlets.

[0032] Compared with experimental groups 1-3, control group 1 showed faster leaf growth in the early stage of transplanting, with leaves starting to grow around 10 days. However, some plants began to rot at the rootstock starting around 15 days later. Figure 2 After 30 days, all 20 peony plants died. The results showed that in control group 1, the leaves of the 20 peony plants unfolded earlier but the root system grew slowly. The plants could not receive timely nutrient supply, resulting in weak growth. The pathogens multiplied rapidly, and the plants eventually died from pathogen infection.

[0033] In summary, compared with existing transplanting environments, this invention overcomes the shortcomings of traditional cultivation techniques, such as poor root growth, long recovery time, and susceptibility to pathogens. This invention ensures that the root system of peony test-tube seedlings recovers rapidly, shortens the recovery time, and achieves successful transplanting and cultivation of peony test-tube seedlings. The survival rate of the 20 peony plants in experimental group 2 was as high as 35%, which is of great significance for the large-scale cultivation of peony test-tube seedlings.

[0034] Example 2: Effect of transplanting substrate moisture content on the survival rate of peony test-tube seedlings after transplanting

[0035] The first step is to mix peat moss, perlite, and vermiculite in a volume ratio of 2:2:1, and then mix them evenly with tap water to obtain transplanting substrates with different moisture contents.

[0036] The second step is to fill the transplanting substrate into the nutrient pots, transplant the peony test-tube seedlings into each nutrient pot, and place the peony plants in a constant temperature plant incubator.

[0037] In this embodiment, the peony test-tube seedlings were divided into four groups according to the moisture content of the transplanting substrate, with the moisture content of the transplanting substrate being 56-60%, 61-65%, 66-70%, 71-75%, and ≥76%, respectively.

[0038] The third step is to manage the transplanted peony test-tube seedlings. During this process, the temperature inside the plant culture box is controlled at 15℃ (i.e., the peony test-tube seedlings are kept at a constant temperature of 15℃). For the first 15 days after transplanting, the seedlings are covered to maintain moisture. After 15 days, the covers are gradually opened for ventilation. In addition, the moisture content of the transplanting substrate is controlled by the amount and frequency of watering. The watering amounts for the four groups of peony test-tube seedlings are 10mL / plant (corresponding to the 56-60% group), 10-30mL / plant (corresponding to the 61-65% group), 30-50mL / plant (corresponding to the 66%-70% group), 50-100mL / plant (corresponding to the 71%-75% group), and 100mL / plant (corresponding to the ≥76% group), respectively, to ensure that the moisture content of the cultivation substrate for each group remains within the set range.

[0039] Sixty days after planting, the survival rate of each group of peony seedlings in vitro was calculated using conventional methods. The results are shown in Table 2.

[0040] Table 2. Survival rate of four groups of peony plants in Example 2 of the present invention.

[0041]

[0042] The moisture content of the transplanting substrate is an important factor in ensuring plant growth. Appropriate humidity can promote the root growth of peony test-tube seedlings. Excessive substrate moisture will cause the roots of peony test-tube seedlings to rot, while too dry substrate moisture will cause the root tips of peony test-tube seedlings to turn brown, resulting in a low survival rate of peony test-tube seedlings after transplanting.

[0043] Table 2 shows that during post-planting management, under an ambient temperature of 15℃, when the moisture content of the transplanting substrate was within the range of 56%–70%, the survival rate of peony seedlings gradually increased with increasing substrate moisture content. Specifically, when the substrate moisture content was between 61% and 75%, the survival rate of peony seedlings was above 40%. However, when the substrate moisture content was 76%, the survival rate of peony seedlings was 0 after 60 days of post-planting management. The results indicate that the moisture content of the transplanting substrate is easily controlled below 75% during post-planting management of peony seedlings.

[0044] In summary, a moisture content of 56%–75% in the transplanting substrate can promote the survival rate of peony test-tube seedlings after transplanting. Among these, a moisture content of 61%–75% is preferred, and a moisture content of 66%–70% is optimal.

[0045] Example 3: Effect of microbial preparations on the survival rate of peony test-tube seedlings after transplanting

[0046] The first step involves mixing peat moss, perlite, and vermiculite in a volume ratio of 2:2:1, and then mixing them separately with tap water and a microbial preparation to obtain the transplanting substrate. The microbial preparation includes two types: Bacillus subtilis Pro1A2 (purchased from the Horticulture Research Institute of Henan Academy of Agricultural Sciences) and commercially available microbial fertilizer (preferably Jufengchun microbial seedling liquid). The effective viable count of Bacillus subtilis Pro1A2 is 0.8–1.2 billion / mL; the effective bacteria in the microbial fertilizer is Bacillus subtilis, with an effective viable count ≥2.0 billion / mL.

[0047] The above transplanting substrates were divided into three groups: treatment group 1, which was mixed with tap water; treatment group 2, which was mixed with Bacillus subtilis Pro1A2; and treatment group 3, which was mixed with commercially available microbial fertilizer.

[0048] The second step involved filling each group of transplanting substrate into different nutrient pots and transplanting the peony test-tube seedlings into the nutrient pots. The number of seedlings transplanted in treatment groups 1, 2, and 3 was 20 each. Each group of peony plants was then placed in a constant-temperature plant incubator.

[0049] The third step is to manage the transplanted peony seedlings after transplanting. The ambient temperature during the post-planting management process is controlled at 15℃ (that is, the peony plants are placed in an environment of constant temperature of 15℃ for growth).

[0050] During post-planting management, regularly measure the moisture content of the transplanting substrate and add water as needed to ensure that the moisture content of the transplanting substrate is always maintained at 66-70%. In addition, for the first 15 days after planting, cover the substrate to retain moisture, and then open the cover for ventilation.

[0051] During post-planting management, the growth of each group of peony seedlings in vitro was statistically analyzed using conventional methods. The survival rate and growth of the seedlings were assessed 60 days after transplanting. Table 3 shows the survival rate of the peony seedlings 60 days after planting, Table 4 shows the above-ground growth of the seedlings 60 days after planting, and Table 5 shows the overall growth of the seedlings 60 days after planting. Figure 3 .

[0052] Table 3. Effects of microbial agents on the survival rate of peony test-tube seedlings after transplanting.

[0053]

[0054] As shown in Table 3, the survival rate of transplanting substrate mixed with tap water was 50%. The survival rates of treatment groups 2 and 3 treated with microbial agents were higher than those of the tap water treatment group. Among them, the survival rate of treatment group 2, which added Bacillus subtilis Pro1A2, was as high as 80%.

[0055] Table 4. Effects of microbial agents on the aboveground growth of transplanted peony seedlings

[0056]

[0057] Table 4 shows the growth of leaves and petioles of each peony plant group 60 days after planting. Table 4 also shows that tap water, Bacillus subtilis Pro1A2, and microbial fertilizer treatments all promoted the aboveground growth of peony seedlings in vitro. Among these, the transplanted seedlings treated with microbial agents had larger leaves, longer petioles, and a lower leaf length-to-width ratio, exhibiting vigorous growth. In particular, treatment group 2, corresponding to Bacillus subtilis Pro1A2, had an average of 2.1 leaves, an average petiole length of 5.1 cm, and a leaf length-to-width ratio of 0.61.

[0058] Combination Figure 3 It can be seen that the number of leaves and petioles of the peony plants transplanted using the transplanting method of the present invention (i.e., treatment group 2) is much greater than that of the tap water group and the microbial fertilizer group.

[0059] In summary, all three experimental groups promoted the rooting and leaf growth of peony in vitro seedlings, as well as the growth of the above-ground parts, achieving successful transplantation of peony in vitro seedlings. Furthermore, the Bacillus subtilis Pro1A2 treatment group showed the best effect among the three treatment groups, laying the foundation for the large-scale cultivation of peony in vitro seedlings.

Claims

1. A method for improving the survival rate of peony test-tube seedlings after transplanting, characterized in that: The method involves transplanting peony test-tube seedlings into nutrient pots filled with a mixed substrate of peat moss, perlite, and vermiculite. Post-transplanting management is then implemented. The volume ratio of peat moss, perlite, and vermiculite in the transplanting substrate is 2:2:

1. During post-transplanting management, the relative moisture content of the transplanting substrate is maintained at 66%–70%, and the ambient temperature is controlled at 15°C. Initially, the substrate is kept sealed and moisturized; ventilation management begins 12–18 days after transplanting. The transplanting substrate also contains a microbial agent, which is a Bacillus subtilis inoculant. The effective viable count of the Bacillus subtilis inoculant is 0.8 to 1.2 billion / mL, and the effective bacteria are Bacillus subtilis Pro1A2.

2. The method for improving the survival rate of peony test-tube seedlings according to claim 1, characterized in that: The preparation steps of the transplanting substrate include: mixing peat moss, perlite and vermiculite in a volume ratio of 2:2:1, and then mixing them with a microbial agent.

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