Salt-tolerant reed seedling and cultivation method thereof

Abstract

The invention discloses a salt-tolerant reed seedling and a cultivation method thereof. The method comprises the following steps: seed selection and pretreatment; performing exogenous induction treatment; performing germination culture; salt gradient domestication culture; salt tolerance screening, seedling hardening and transplanting. According to the invention, by combining exogenous induction and salt gradient domestication culture, the stress resistance of the plant is regulated, and the salt tolerance mechanism in the seed is activated, so that a foundation is laid for better adaptation and growth in a high-salt environment subsequently; the salt-tolerant reed seedlings induced and domesticated by the method are high in seedling rate, the survival rate of acclimatization and transplantation is 90% or above, the breeding cycle is shortened to be within 100 days, the cultivation cycle is short, and the method is suitable for large-scale popularization and application.

Description

Technical Field

[0001] The present invention relates to the technical field of plant breeding, and particularly to a salt-tolerant reed seedling and a cultivation method thereof. Background Art

[0002] Reed (Phragmites australis) is a perennial herb widely distributed in wetlands and saline-alkali lands, with important ecological and economic values. However, the tolerance of common reed to high-salt environments is limited, which restricts its application in saline-alkali lands.

[0003] Traditional cultivation methods for salt-tolerant plants mainly rely on natural selection or genetic engineering, but they have problems such as long cycle, high cost, and complex technology. Summary of the Invention

[0004] The present invention solves the above-mentioned deficiencies in the cultivation of existing salt-tolerant reeds by providing a salt-tolerant reed seedling and a cultivation method thereof.

[0005] To solve the above technical problems, the present invention provides a cultivation method for salt-tolerant reed seedlings, comprising the following steps:

[0006] (1) Seed selection and pretreatment: Select healthy and plump reed seeds, and perform soaking and disinfection treatments;

[0007] (2) Exogenous induction treatment: Immerse the disinfected reed seeds in a solution containing an exogenous inducer for 6 - 12 hours; the exogenous inducer is a mixed solution of 100 - 200 mg / L NaCl and 50 - 100 mg / L SA;

[0008] (3) Germination culture: Inoculate the reed seeds after the above exogenous induction treatment into a germination medium and culture for 2 - 3 weeks until the seeds germinate;

[0009] (4) Salt gradient acclimation culture: Transfer the germinated reed seedlings to acclimation media with gradually increasing salt concentrations for salt gradient acclimation culture, and the culture time in each acclimation medium is the same;

[0010] (5) Salt tolerance screening: After the acclimation is completed, transfer the reed seedlings to a salt tolerance screening medium for culture, and screen out salt-tolerant plants with strong growth and well-developed roots;

[0011] (6) Hardening-off and transplantation: First transfer the screened reed seedlings to room temperature for hardening-off for 1 - 2 weeks, and then transplant them to saline-alkali lands or simulated saline-alkali environments.

[0012] In a preferred embodiment of the present invention, in step (3), the germination medium is 1 / 2MS basal medium + 0.5 - 1 mg / L GA3.

[0013] In a preferred embodiment of the present invention, in step (3), the conditions for germination culture are as follows: temperature 25±2°C, light intensity 2000 - 3000 lux, and light time 12 h / day.

[0014] In a preferred embodiment of the present invention, in step (4), the salt concentrations of the acclimation culture medium are successively: 50 mM, 100 mM, 150 mM, 200 mM; and the culture time in each acclimation culture medium is 2 - 3 weeks.

[0015] In a preferred embodiment of the present invention, in step (5), the salt - tolerance screening culture medium is 1 / 2MS basal medium + 200 mM NaCl.

[0016] In a preferred embodiment of the present invention, in step (5), the conditions for seedling hardening are as follows: greenhouse environment, humidity 60% - 70%, and light intensity 5000 - 6000 lux.

[0017] The beneficial effects of the present invention are as follows: The present invention provides a salt - tolerant reed seedling and its cultivation method. By combining exogenous induction and salt - gradient acclimation culture, it regulates plant stress resistance and activates the internal salt - tolerance mechanism of seeds, laying a foundation for better adaptation and growth in high - salt environments in the future; the salt - tolerant reed seedlings induced and acclimated by the present invention have a high seedling formation rate, the survival rate of seedling hardening and transplantation is above 90%, and the propagation cycle is shortened to within 100 days. The cultivation cycle is short, making it suitable for large - scale popularization and application. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] Figure 1 It is a schematic diagram of the process of a cultivation method for a salt - tolerant reed seedling of the present invention. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0019] The following elaborates on the preferred embodiments of the present invention in conjunction with the drawings, so that the advantages and features of the present invention can be more easily understood by those skilled in the art, thereby making a clearer and more definite definition of the protection scope of the present invention.

[0020] Example 1

[0021] A cultivation method for a salt - tolerant reed seedling is as follows:

[0022] (1) Seed selection and pretreatment: Select healthy and plump reed seeds, soak them in clean water for 24 hours, and remove the unplump seeds floating on the water surface; then disinfect the remaining seeds with 0.1% sodium hypochlorite solution for 10 minutes, and then rinse them with sterile water 3 - 5 times to complete the disinfection pretreatment.

[0023] (2) Exogenous induction treatment: The disinfected reed seeds were immersed in a solution containing an exogenous inducer for 8 h. The exogenous inducer used was a mixed solution of 150 mg / L NaCl and 75 mg / L SA (salicylic acid). After the immersion, the reed seeds were taken out and rinsed thoroughly with sterile water.

[0024] The exogenous inducer can enhance the salt tolerance and stress resistance of reed seeds through chemical signal regulation, making preparations for adaptive domestication, and thus helping the seeds to better adapt and grow in the subsequent high-salt environment. It specifically includes the following aspects:

[0025] Improving salt tolerance: Low-concentration NaCl in the exogenous inducer can simulate a salt stress environment during the induction stage, activate the salt tolerance mechanism inside the seeds, and thus induce the seeds to produce adaptive responses, such as accumulating osmotic adjustment substances (such as proline, soluble sugars, etc.) and antioxidant enzymes (such as superoxide dismutase, peroxidase, etc.), to improve the tolerance of the seeds in the subsequent high-salt environment.

[0026] Enhancing stress resistance: SA has the function of regulating the stress resistance response of plants. It can activate the defense mechanism of plants through signal transduction pathways, such as inducing the activity of antioxidant enzymes and reducing the accumulation of reactive oxygen species (ROS), thereby reducing the damage of salt stress to cells.

[0027] Promoting seed germination and seedling growth: The treatment with the exogenous inducer can improve the physiological state of the seeds and promote seed germination and early seedling growth. By activating the metabolic pathways inside the seeds, the inducer can provide better energy and material bases for the seeds, making them show stronger growth potential during the subsequent germination and domestication processes.

[0028] (3) Germination culture: The reed seeds after the above exogenous induction treatment were inoculated into a germination medium and cultured for 2 weeks under the conditions of a temperature of 25 ± 2 °C, a light intensity of 2500 lux, and a light time of 12 h / day to germinate the reed seeds;

[0029] Specifically, the germination medium was 1 / 2 MS basal medium + 0.8 mg / L GA3 (gibberellin 3).

[0030] (4) Salt gradient domestication culture: The germinated reed seedlings were first transferred to a 1 / 2 MS basal medium with an NaCl concentration of 50 mM and cultured for 2 weeks under the conditions of a temperature of 25 ± 2 °C, a light intensity of 2500 lux, and a light time of 12 h / day;

[0031] Then, the salt-tolerant plants (plants with good growth status) were screened out and transferred to a 1 / 2 MS basal medium with an NaCl concentration of 100 mM and cultured for 2 weeks under the conditions of a temperature of 25 ± 2 °C, a light intensity of 2500 lux, and a light time of 12 h / day;

[0032] Transfer the (plants with good growth status) to a 1 / 2MS basal medium with a NaCl concentration of 150 mM, and culture for 2 weeks under the conditions of a temperature of 25 ± 2°C, a light intensity of 2500 lux, and a light duration of 12 h / day;

[0033] Finally, transfer the (plants with good growth status) to a 1 / 2MS basal medium with a NaCl concentration of 200 mM, and culture for 2 weeks under the conditions of a temperature of 25 ± 2°C, a light intensity of 2500 lux, and a light duration of 12 h / day;

[0034] After 8 weeks of acclimation culture in the above four salt gradient media, gradually induce the expression of salt-tolerant genes in reed seeds, thereby inducing and obtaining salt-tolerant plants.

[0035] (5) Salt tolerance screening: After the acclimation, transfer the reed seedlings to a salt tolerance screening medium, and culture for 2 weeks under the conditions of a temperature of 25 ± 2°C, a light intensity of 2500 lux, and a light duration of 12 h / day, and screen out salt-tolerant plants with strong growth and well-developed roots;

[0036] Specifically, the salt tolerance screening medium is 1 / 2MS basal medium + 200 mM NaCl.

[0037] (6) Hardening off and transplanting: First, transfer the screened reed seedlings to room temperature for hardening off, gradually reduce the humidity to 60%-70%, and increase the light intensity to 5000 lux. The hardening off period is 2 weeks.

[0038] Through hardening off, gradually adjust the environmental conditions to enable the reed seedlings to smoothly transition from the artificial culture environment to the natural environment, enhance their environmental adaptability, stress resistance, and physiological functions, and ultimately improve the transplanting survival rate and growth performance in saline-alkali land.

[0039] After the hardening off, transplant the reed seedlings to saline-alkali land or a simulated alkali environment for routine management, and the seedling survival rate reaches over 90%.

[0040] Example 2

[0041] A method for cultivating salt-tolerant reed seedlings, the specific steps are as follows:

[0042] (1) Seed selection and pretreatment: Select healthy and plump reed seeds, soak them in clean water for 24 hours, and remove the unplump seeds floating on the water surface; then disinfect the remaining seeds with a 0.1% sodium hypochlorite solution for 10 minutes, and rinse with sterile water 3-5 times to complete the disinfection pretreatment.

[0043] (2) Exogenous induction treatment: The disinfected reed seeds were immersed in a solution containing an exogenous inducer for 12 h. The exogenous inducer used was a mixed solution of 200 mg / L NaCl and 100 mg / L SA (salicylic acid). After the immersion, the reed seeds were taken out and rinsed thoroughly with sterile water.

[0044] (3) Germination culture: The reed seeds after the above exogenous induction treatment were inoculated into a germination medium and cultured for 3 weeks under the conditions of a temperature of 25 ± 2 °C, a light intensity of 2000 lux, and a light duration of 12 h / day to germinate the reed seeds;

[0045] Specifically, the germination medium was 1 / 2MS basal medium + 1.0 mg / L GA3 (gibberellin 3).

[0046] (4) Salt gradient acclimation culture: The germinated reed seedlings were first transferred to a 1 / 2MS basal medium with an NaCl concentration of 50 mM and cultured for 2 weeks under the conditions of a temperature of 25 ± 2 °C, a light intensity of 2500 lux, and a light duration of 12 h / day;

[0047] Then, the salt-tolerant plants (plants with good growth status) were selected and transferred to a 1 / 2MS basal medium with an NaCl concentration of 100 mM and cultured for 2 weeks under the conditions of a temperature of 25 ± 2 °C, a light intensity of 2500 lux, and a light duration of 12 h / day;

[0048] Next (plants with good growth status) were transferred to a 1 / 2MS basal medium with an NaCl concentration of 150 mM and cultured for 2 weeks under the conditions of a temperature of 25 ± 2 °C, a light intensity of 2500 lux, and a light duration of 12 h / day;

[0049] Finally (plants with good growth status) were transferred to a 1 / 2MS basal medium with an NaCl concentration of 200 mM and cultured for 2 weeks under the conditions of a temperature of 25 ± 2 °C, a light intensity of 2500 lux, and a light duration of 12 h / day;

[0050] After 8 weeks of acclimation culture in the above four salt gradient media, the expression of salt-tolerant genes in the reed seeds was gradually induced, thereby inducing and obtaining salt-tolerant plants.

[0051] (5) Salt tolerance screening: After the acclimation, the reed seedlings were transferred to a salt tolerance screening medium and cultured for 2 weeks under the conditions of a temperature of 25 ± 2 °C, a light intensity of 2500 lux, and a light duration of 12 h / day to screen out salt-tolerant plants with strong growth and well-developed roots;

[0052] Specifically, the salt tolerance screening medium was 1 / 2MS basal medium + 200 mM NaCl.

[0053] (6) Hardening off and transplanting: Transplant the selected reed seedlings to room temperature for hardening off, gradually reduce the humidity to 60%-70%, and increase the light intensity to 5000 lux. The hardening-off period is 2 weeks.

[0054] After the hardening-off is completed, transplant the reed seedlings to saline-alkali land or a simulated alkali environment for routine management. The survival rate of the seedlings is over 93%.

[0055] Comparative Example 1

[0056] Compared with Example 1, the difference is that after the selected seeds are pretreated, they are not subjected to exogenous induction treatment, but directly inoculated in the germination medium for cultivation, and then subjected to salt gradient acclimation cultivation, salt tolerance screening, and hardening off in sequence. The treatment conditions are the same as those in Example 1.

[0057] After the hardening-off is completed, transplant the reed seedlings to saline-alkali land or a simulated alkali environment for routine management. The survival rate of the seedlings is less than 80%.

[0058] Comparative Example 2

[0059] Compared with Example 1, the difference is that after the selected reed seeds are disinfected, subjected to exogenous induction treatment, germination cultivation, salt gradient acclimation cultivation, and salt tolerance screening, they are directly transplanted to saline-alkali land or a simulated alkali environment for routine management without going through the hardening-off process. The survival rate of the seedlings is 87%.

[0060] The method for inducing and domesticating the cultivation of salt-tolerant reed seedlings in the present invention combines exogenous induction and salt gradient acclimation cultivation. SA is used to regulate the plant stress response. A low concentration of NaCl can simulate a salt stress environment during the induction stage, activate the salt tolerance mechanism inside the seeds, and the two form a synergistic effect, enhancing the salt tolerance and stress resistance of reed seeds, laying a foundation for better adaptation and growth in a high-salt environment in the future.

[0061] The salt-tolerant reed seedlings induced and domesticated by the present invention have a high seedling formation rate and a short cultivation period. The cultivated salt-tolerant reed seedlings can be used for the ecological restoration and resource utilization of saline-alkali land, and have important ecological and economic values.

[0062] The above are only the embodiments of the present invention, and do not limit the patent scope of the present invention accordingly. Any equivalent structure or equivalent process transformation made by using the content of the specification and drawings of the present invention, or directly or indirectly applied to other related technical fields, shall be included in the patent protection scope of the present invention by the same token.

Claims

1. A method for cultivating salt-tolerant reed seedlings, characterized in that: The steps include: (1) Seed selection and pretreatment: Select healthy and plump reed seeds, soak them and disinfect them; (2) Exogenous induction treatment: the sterilized reed seeds were placed in a solution containing an exogenous inducer and soaked for 6-12 hours; the exogenous inducer was a mixed solution of 100-200 mg / L NaCl and 50-100 mg / L SA; (3) Germination culture: inoculating the reed seeds treated with the above exogenous induction into a germination medium and culturing for 2-3 weeks until the seeds germinate; (4) Salt gradient acclimation culture: after germination, the reed seedlings are transferred to acclimation culture medium with increasing salt concentrations for salt gradient acclimation culture, and the culturing time in each acclimation culture medium is the same; (5) Salt tolerance screening: After the acclimatization, the reed seedlings are transferred to the salt tolerance screening medium for cultivation to screen out salt-tolerant plants with strong growth and well-developed root systems; (6) Hardening and transplanting: The selected reed seedlings are first moved to room temperature for hardening for 1-2 weeks, and then transplanted to saline-alkali land or simulated saline-alkali environment.

2. The method for cultivating salt-tolerant reed seedlings according to claim 1, characterized in that: In step (3), the germination medium is 1 / 2MS basal medium + 0.5-1 mg / L GA3.

3. The method for cultivating salt-tolerant reed seedlings according to claim 1, characterized in that: In step (3), the conditions for the germination culture are: temperature 25±2°C, light intensity 2000-3000 lux, and illumination time 12h / day.

4. The method for cultivating salt-tolerant reed seedlings according to claim 1, characterized in that: In step (4), the salt concentrations of the acclimation culture medium are: 50mM, 100mM, 150mM, 200mM, respectively; and the culturing time in each acclimation culture medium is 2-3 weeks.

5. The method for cultivating salt-tolerant reed seedlings according to claim 1, characterized in that: In step (5), the salt-tolerance screening medium is 1 / 2MS basal medium + 200 mM NaCl.

6. The method for cultivating salt-tolerant reed seedlings according to claim 1, characterized in that: In step (5), the conditions for hardening the seedlings are: a greenhouse environment, a humidity of 60%-70%, and a light intensity of 5000-6000 lux.

7. A salt-tolerant reed seedling, characterized in that: The method according to any one of claims 1 to 6 is used to culture the obtained product.

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