Safe idesia seed germination induction method

Abstract

The invention discloses a safe idesia seed germination induction method. The method comprises the following steps: (1) removing wax on the surfaces of idesia seeds, washing and draining to obtain wax-removed idesia seeds; (2) putting the idesia seeds subjected to wax removal into a gibberellin GA3 solution with the concentration of 0-200mg / L, shaking and soaking for 24-36 hours, washing and draining to obtain the idesia seeds treated by GA3; (3) after sterilizing and disinfecting the idesia seeds, transferring the idesia seeds into a metaphosphoric acid aqueous solution with the mass concentration of 10-50%, shaking and soaking for 25-45 minutes, carrying out germination promoting treatment, washing and draining to obtain the idesia seeds subjected to germination promoting treatment; and (4) spreading the idesia polycarpa seeds subjected to germination promoting treatment into a germination induction culture medium for germination induction. According to the method, a controlled dangerous chemical reagent is not used, new or old seeds are not picked out, the seeds do not need to be laminated, the seeds can germinate in 5 days and enter a germination peak in 15 days, the germination rate can reach 92.59%, and the method is an extremely efficient and safe idesia polycarpa seed germination induction method.

Description

Technical Field

[0001] The invention relates to the technical field of seed germination, in particular to a safe method for inducing germination of Castanopsis chinensis seeds. Background Art

[0002] Idesia polycarpa Maxim. is a perennial deciduous tree of the genus Idesia in the Salicaceae family. It has outstanding value in ecological protection, gardening and economic development due to its fast growth and strong resistance to adversity, bright fruit color and long fruiting period, high fruit yield and high oil content, and the demand for seedlings continues to grow.

[0003] At present, the propagation of Castanopsis chinensis seedlings is mainly based on seed propagation. However, because the seed coat of Castanopsis chinensis seeds is hard, covered with wax, and rich in germination inhibitors, its dormancy period is long and difficult to break, making it difficult to germinate naturally. Under field and nursery conditions, the germination rate is only about 20%, and it takes more than 60 days to enter the initial germination stage. Low temperature (5℃-15℃) stratification assisted sowing can increase the germination rate to about 51% and shorten the initial germination time to about 20 days. The use of concentrated sulfuric acid and hydrogen peroxide can further improve the germination rate of Castanopsis chinensis seeds, with a significant germination-promoting effect, which can increase the germination rate to about 90% (Shen Yangcheng, Hou Jinyan, Liu Wenbo, Jin Shan, Wu Lifang. 2015. Research on rapid and efficient germination of Castanopsis chinensis. Chinese Agricultural Science Bulletin, 31(13):5-9; CN103814654A). However, hydrogen peroxide and concentrated sulfuric acid are both highly corrosive hazardous chemicals and are safety-controlled reagents in my country. Their procurement and use must be reviewed by the public security department. Only units / enterprises with corresponding qualifications (including professional hazardous chemical transportation, storage, use, waste liquid recovery and process monitoring systems, etc.) can apply for their use. They are inconvenient to use and pose a major social safety risk.

[0004] In recent years, a patent application with publication number CN115843693A has published a method for rapid germination of Castanopsis chinensis seeds. The method involves soaking the seeds with 200 mg / L gibberellin GA7 and using a solution containing 11 mg / L GA7. 4+7 The culture medium promotes germination, which can make the germination rate of Castanopsis chinensis seeds reach 89%. This method avoids the dependence on concentrated sulfuric acid / hydrogen peroxide when artificially inducing Castanopsis chinensis to germinate and break its lengthy dormancy. However, the above methods all have strict requirements on the age of seeds, and all need to use Castanopsis chinensis seeds newly harvested that year, and cannot use old seeds. Usually, old Castanopsis chinensis seeds have difficulty germinating and can only be discarded and cannot be used for seedling cultivation. This is a huge waste of resources for Castanopsis chinensis, which is famous for its high yield (fruit yield / seed quantity). In fact, new and old seeds are mixed in the Castanopsis chinensis seed sales market and cannot be distinguished by the naked eye. Therefore, the Castanopsis chinensis seeds purchased by users are often a mixture of new and old seeds, which greatly reduces the emergence rate of Castanopsis chinensis seedlings in production. Summary of the Invention

[0005] In view of this, the present invention proposes a method for inducing germination of Castanopsis schefflera seeds, which is simple and safe to operate, has high germination efficiency, and is not picky about seed age.

[0006] The present invention provides a method for inducing germination of Castanopsis truncatula seeds, comprising the following steps:

[0007] (1) removing wax from the surface of the Castanopsis truncatula seeds, washing them and filtering them to obtain wax-removed Castanopsis truncatula seeds;

[0008] (2) placing the tung oil seed after wax removal in step (1) in a 0-200 mg / L gibberellin GA3 solution, shaking and soaking for 24-36 hours to ensure that the seeds absorb sufficient water to assist in disrupting dormancy; washing and filtering to obtain GA3-treated tung oil seeds;

[0009] (3) sterilizing the GA3-treated Castanopsis chinensis seeds in step (2), placing them in a metaphosphoric acid aqueous solution with a mass concentration of 10% to 50%, shaking and soaking them for 25 to 45 minutes, and performing a germination-promoting treatment to enhance seed coat permeability and interfere with intracellular signal transduction through the interaction between metaphosphoric acid and seed protein, thereby breaking seed dormancy; washing and filtering to obtain Castanopsis chinensis seeds after germination-promoting treatment;

[0010] (4) Sowing the Castanopsis chinensis seeds after the germination promotion treatment in step (3) into a germination induction medium for germination induction.

[0011] Preferably, the mass concentration of the metaphosphoric acid aqueous solution in step (3) is 10% to 30%.

[0012] Preferably, the step (3) is performed by shaking and soaking in the metaphosphoric acid aqueous solution for 25 to 35 minutes.

[0013] Preferably, the germination induction medium in step (4) is based on 1 / 2MS, supplemented with 0.2 mg / L to 1.0 mg / L of 6-BA, 0.1 mg / L to 0.3 mg / L of IBA, 25 g / L to 30 g / L of sucrose, and 7.0 g / L to 8.0 g / L of carrageenan; and the pH value of the germination induction medium is 5.8 to 6.0.

[0014] Further preferably, the germination induction medium is based on 1 / 2MS, supplemented with 1.0 mg / L 6-BA, 0.3 mg / L IBA, 25 g / L sucrose and 7.0 g / L carrageenan.

[0015] Preferably, the germination induction in step (4) is to culture the Castanopsis schrenkiana seeds under conditions of a temperature of 25°C to 28°C, a light intensity of 1200 to 1800 Lx, and a photoperiod of 14 hours light / 10 hours dark for 10 to 60 days.

[0016] Further preferably, the germination induction is to culture the Castanopsis schrenkiana seeds under conditions of a temperature of 25°C, a light intensity of 1200 Lx, and a photoperiod of 14 hours of light / 10 hours of darkness for 30 days.

[0017] Preferably, the method for removing the wax on the surface of the Castanopsis schrenkiana seeds in step (1) is: placing the Castanopsis schrenkiana seeds in a cloth bag, adding detergent, rubbing and rinsing repeatedly until the foam is not yellow, so as to fully remove the wax on the surface of the Castanopsis schrenkiana seeds.

[0018] Preferably, the sterilization method in step (3) is: soaking the Castanopsis schrenkiana seeds treated with gibberellins in 75% alcohol for 30 to 40 seconds, and rinsing them with sterile water 3 to 4 times; soaking them in a sodium hypochlorite solution with an effective chlorine concentration of 6% to 8% for 30 to 40 minutes, and rinsing them with sterile water 3 to 4 times to completely disinfect the Castanopsis schrenkiana seeds.

[0019] Preferably, the Castanopsis aviculare seeds are new seeds or aged seeds with a storage period of ≤8 years.

[0020] The present invention provides an efficient germination induction method for Castanopsis chinensis seeds, which includes three steps: seed disinfection, germination promotion treatment, and germination induction. This method can enable Castanopsis chinensis seeds to germinate within 5 days without stratification, enter the peak germination period on the 15th day, and the germination rate can exceed 90% on the 30th day; it also has a germination-promoting effect on old Castanopsis chinensis seeds with a storage period of ≤8 years. This method shortens the production cycle of Castanopsis chinensis seedlings, improves the seed germination rate, enhances resource utilization efficiency, and greatly reduces the production risks caused by unknown seed age. At the same time, the method provided by the present invention does not use safety-controlled hazardous chemicals, and the procurement / supervision of chemicals is convenient, which greatly improves production safety. The Castanopsis chinensis seeds used in this method do not require stratification and are not picky about new / old seeds. The seeds can germinate within 5 days and enter the peak germination period on the 15th day, with a germination rate of 92.59%. It is an extremely efficient Castanopsis chinensis seedling cultivation method. In addition, the safe and efficient seed germination method provided by the present invention can be applied to the propagation of Castanopsis schrenkiana tissue culture seedlings under sterile conditions, laying a technical foundation for the later expansion and propagation of Castanopsis schrenkiana seedlings, genetic modification and creation of new species. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] For purposes of illustration and not limitation, the present invention will now be described with reference to preferred embodiments thereof, particularly with reference to the accompanying drawings, in which:

[0022] Figure 1 These are the appearance pictures of fresh fruits and aged seeds of Castanopsis chinensis; among them: A is the appearance picture of Castanopsis chinensis fruits; B is the appearance picture of 2-year-old Castanopsis chinensis seeds.

[0023] Figure 2The germination effect of Castanopsis chinensis seeds after 12 days of combined treatment with gibberellin (GA3) and concentrated sulfuric acid / hydrogen peroxide; among them: A is the germination status of seeds soaked in 0 mg / L gibberellin combined with concentrated sulfuric acid treatment (30%, 20 min); B is the germination status of seeds soaked in 100 mg / L gibberellin combined with concentrated sulfuric acid treatment (30%, 20 min); C is the germination status of seeds soaked in 200 mg / L gibberellin combined with concentrated sulfuric acid treatment (30%, 20 min); D is the germination status of seeds soaked in 0 mg / L gibberellin combined with hydrogen peroxide treatment (30%, 20 min); E is the germination status of seeds soaked in 100 mg / L gibberellin combined with hydrogen peroxide treatment (30%, 20 min); F is the germination status of seeds soaked in 200 mg / L gibberellin combined with hydrogen peroxide treatment (30%, 20 min); the culture medium used was M6 and the seeds were all 2 years old.

[0024] Figure 3 The comparison of early (5d) germination differences of Aglaonema tung seeds treated with different reagents; among them: A is 30% hydrogen peroxide treatment for 20 minutes; B is 30% concentrated sulfuric acid treatment for 20 minutes; C is 20% metaphosphoric acid treatment for 30 minutes.

[0025] Figure 4 The germination-promoting effect of 20% metaphosphoric acid on aged Castanopsis truncatula seeds; among them: A is 8-year-old seed; B is 4-year-old seed; C is 2-year-old seed.

[0026] Figure 5 This is a comparison of the germination-promoting effects of metaphosphoric acid (red) and concentrated sulfuric acid (green) on Castanopsis chinensis seeds; among them: the metaphosphoric acid (red broken line) treatment method is 20% metaphosphoric acid treatment for 30 minutes; the concentrated sulfuric acid (green broken line) treatment method is 30% concentrated sulfuric acid treatment for 20 minutes. DETAILED DESCRIPTION

[0027] Unless otherwise specified, the experimental methods used in the following examples are conventional methods.

[0028] Unless otherwise specified, the materials and reagents used in the following examples can be obtained from commercial sources.

[0029] Example 1: Optimization of the germination induction medium for Castanopsis chinensis seeds

[0030] Take 2-year-old tung oil seed ( Figure 1The non-patent literature that records the seeds of the Chinese tallow tree is: Li Dawei, Jiang Xibing, Wu Kaiyun, Peng Jialong, Bai Jianjie. 2012. Analysis of fatty acid composition variation of Chinese tallow tree fruits from different provenances. Acta Physiologica Sinica. 48(5):505-510). Rub and rinse repeatedly with detergent until the foam is washed out without yellow color, ensuring that the surface wax is fully removed. Thereafter, rinse with running water and drain to obtain the Chinese tallow tree seeds after wax removal. Soak the Chinese tallow tree seeds after wax removal in 0-200 mg / L gibberellin (GA3) solution, place them on a shaking table with an oscillation frequency of 200 r / min and shake for 24-36 hours to ensure that the seeds absorb enough water to help disrupt dormancy. Then, wash the seeds with clean water and drain to obtain the Chinese tallow tree seeds treated with gibberellin. Transfer them to sterile tissue culture bottles and perform subsequent operations in a sterilized clean bench.

[0031] After gibberellin treatment, Castanopsis chinensis seeds were soaked in 75% (v / v) alcohol for 30 seconds, rinsed three times with sterile water, and shaken for 3-5 minutes each time. After draining, the seeds were soaked in sodium hypochlorite with an available chlorine concentration of 8% by mass for 30 minutes, rinsed three to five times with sterile water, and shaken for 8-10 minutes each time. After draining, the seeds were subjected to traditional germination promotion treatment to compare whether there was any difference in response between the two traditional methods: the seeds were soaked in 30% (v / v) concentrated sulfuric acid or 30% (v / v) hydrogen peroxide for 20 minutes, rinsed five to six times with sterile water, and shaken for 10-15 minutes each time. After draining, the surface water droplets were removed with sterile filter paper, and the seeds were evenly spread on germination induction medium (Table 1). The culture bottles were placed at 25°C, with a light intensity of 1200 Lx and a photoperiod of 14 hours light / 10 hours dark. The seeds were observed daily for whitening and growth.

[0032] The germination promotion treatment in this embodiment is set up with seven culture medium schemes M1-M7. The application effects of these seven culture medium schemes are shown in Table 1. Figure 2 and Figure 3 shown.

[0033] Table 1 Germination-promoting effect of culture medium on germination induction of Castanopsis chinensis seeds

[0034]

[0035] As shown in Table 1, the old seeds of Castanopsis chinensis treated with hydrogen peroxide and concentrated sulfuric acid are not suitable for germination on the blank MS or 1 / 2MS medium (M1-M2) used for conventional sowing, and require hormone assistance (M4-M7). Among them, NAA is basically ineffective, and low-dose GA3 has a certain germination-promoting effect, but the germination period is long, and it takes more than a month for the seeds to slowly turn white, which is inefficient. In contrast, M6 medium has the best induction effect on the germination of Castanopsis chinensis seeds, which can help the seeds enter the germination peak around the 15th day. Thus, the optimal medium for inducing the germination of Castanopsis chinensis seeds was screened as follows: based on 1 / 2MS, 6-BA 1.0 mg / L, IBA 0.3 mg / L, sucrose 25 g / L and carrageenan 7.0 g / L were added; the pH value of the germination induction medium was 5.8-6.0.

[0036] Figure 2 This paper compares the germination-promoting effects of gibberellin GA3 and concentrated sulfuric acid / hydrogen peroxide on Castanopsis chinensis seeds. Figure 2 It can be seen that the concentrated sulfuric acid in the traditional germination promotion method ( Figure 2 A, B, C) and hydrogen peroxide ( Figure 2 Treatments D, E, and F can promote the germination of old seeds of Castanopsis chinensis, and the effect is better when combined with GA3 seed soaking in advance. Whether it is concentrated sulfuric acid treatment or hydrogen peroxide treatment, adding low-concentration GA3 treatment (100 mg / L, Figure 2 B and E) can significantly promote germination; but further increasing the GA3 concentration to 200 mg / L ( Figure 2 C and F) showed no significant additional benefit on seed germination. Therefore, for the germination of Castanopsis chinensis seeds, a GA3 concentration of no more than 200 mg / L is appropriate; from an economic perspective, 100 mg / L GA3 can be used for seed soaking.

[0037] Figure 3 The results show that the early (5-day) germination differences of Tung oilseed seeds treated with hydrogen peroxide (30% concentration soaked for 20 minutes) and concentrated sulfuric acid (30% concentration soaked for 20 minutes) were compared. Among them, the cotyledon germination of seeds treated with hydrogen peroxide was faster ( Figure 3 A), but the hypocotyl elongation is insufficient, which is not conducive to normal development in the later stage and is prone to deformity; and hydrogen peroxide is not easy to remove, and its residue on the seed coat will hinder seed germination, so the concentration and duration of use must be strictly controlled. Relatively speaking, the germination consistency of Tung oil seed treated with concentrated sulfuric acid is better, and the hypocotyl elongates first ( Figure 3 B) showed normal growth. Therefore, concentrated sulfuric acid treatment was used as a control method to promote germination.

[0038] Example 2: Optimization of the germination-promoting conditions of tung oil seed by metaphosphate

[0039] Take the seeds of Castanopsis chinensis, including old Castanopsis chinensis seeds stored for 2, 4 and 8 years, and rub and rinse them repeatedly with detergent until the foam is washed out without yellow, ensuring that the surface wax is fully removed. After that, rinse with running water and drain to obtain the Castanopsis chinensis seeds after wax removal. Soak the seeds in 100mg / L gibberellin GA3 solution and place them on a shaking table with an oscillation frequency of 200r / min for 24h to ensure that the seeds absorb enough water to assist in disrupting dormancy. Then, wash the seeds with clean water, drain, and obtain the Castanopsis chinensis seeds treated with gibberellin. Transfer them to a sterile tissue culture bottle and perform subsequent germination promotion operations in a disinfected clean bench.

[0040] The seeds of Castanopsis chinensis treated with gibberellin were soaked in 75% alcohol (v / v) for 30 seconds, rinsed with sterile water 3 to 5 times, and shaken and soaked for 3 to 5 minutes each time; after draining, the seeds were soaked in sodium hypochlorite with an effective chlorine concentration of 8% for 30 minutes, rinsed with sterile water 3 to 5 times, and shaken and soaked for 8 to 10 minutes each time; after draining, the seeds were subjected to a germination promotion treatment, that is, the seeds were shaken and soaked in 10% to 50% metaphosphoric acid (w / w) for 25 to 45 minutes, rinsed with sterile water 5 to 6 times, and shaken and soaked for 10 to 15 minutes each time; drained, the surface water droplets were absorbed with sterile filter paper, and the seeds were evenly sprinkled into M6 germination induction medium; the culture bottles were placed under the conditions of a temperature of 25°C, a light intensity of 1200Lx, and a photoperiod of 14 hours of light / 10 hours of darkness for 30 days, and the whitening and growth of the seeds were observed and recorded every day.

[0041] The effects of the sprout promotion treatment in this embodiment are shown in Table 2. Figure 3 and Figure 4 shown.

[0042] Table 2 Effect of metaphosphate treatment on the germination promotion of Castanopsis chinensis seeds

[0043]

[0044] From Table 2, Figure 3 and Figure 4 The results showed that metaphosphate treatment could significantly increase the germination rate of Castanopsis chinensis seeds, and the hypocotyls of the seeds after treatment elongated first during germination ( Figure 3 Middle C), the seeds developed normally, which is similar to the classic concentrated sulfuric acid germination promotion effect ( Figure 3 B); Metaphosphoric acid is a general safety control reagent, and its purchase, storage and operation safety are much higher than concentrated sulfuric acid; and it is still effective for many years of aging ( Figure 4 However, the germination-promoting effect of metaphosphate showed obvious effects of seed age and dosage (Table 2). Among them, the seed storage years had a greater impact on the germination rate than the dosage effect: Figure 4 The germination promoting effect of 20% metaphosphoric acid on aged seeds of Castanopsis chinensis, among which, 2-year-old ( Figure 4 A), 4 years old ( Figure 4 Medium B) and 8 years old ( Figure 4 The germination rate of Castanopsis chinensis seeds decreased from 92.59% to 40.52% and 18.04% in C. Regarding the dose effect (Table 2), the germination rate of slightly aged seeds increased slightly with increasing metaphosphate concentration. For example, treatment of two-year-old Castanopsis chinensis seeds with 10% and 20% metaphosphate increased their germination rate from 89.79% to 92.59%, and further increases in concentration maintained the germination rate. However, for moderately aged seeds (four years old), the germination rate decreased from 40.52%, 35.13%, and 32.56% to 29.67% as the metaphosphate concentration increased from 20%, 30%, 40%, and 50%. Therefore, 20% metaphosphate is the optimal concentration for promoting Castanopsis chinensis seed germination.

[0045] Furthermore, the duration of metaphosphoric acid soaking significantly affected its germination-promoting effect on Castanopsis chinensis seeds (Table 2). The inhibitory effect was more pronounced with older seeds and higher metaphosphoric acid concentrations: Soaking eight-year-old Castanopsis chinensis seeds in 30% metaphosphoric acid for 25, 35, and 45 minutes reduced their germination rate from 14.43% to 12.89% and 12.77%, respectively. However, at lower metaphosphoric acid concentrations, extending the treatment time improved germination rates: Soaking eight-year-old Castanopsis chinensis seeds in 20% metaphosphoric acid for 25 and 35 minutes resulted in slightly higher germination rates of 13.56% and 18.04%, respectively. Therefore, the optimal treatment duration for promoting Castanopsis chinensis seed germination is 25 to 35 minutes.

[0046] Therefore, the optimal method for promoting germination of Castanopsis chinensis seeds was screened as follows: first, repeatedly rub and rinse with detergent to remove the wax on the seed surface, and then soak the seeds in 100 mg / L GA3 solution on a shaker with an oscillation frequency of 200 r / min for 24 hours. After that, transfer to the clean bench, soak the seeds in 75% alcohol for 30 seconds, rinse with sterile water 3 to 5 times, and shake and soak for 3 to 5 minutes each time; after draining, soak the seeds in sodium hypochlorite with 8% effective chlorine for 30 minutes, rinse with sterile water 3 to 5 times, and shake and soak for 8 to 10 minutes each time; after draining, perform germination promotion treatment on the seeds, that is, soak in 20% metaphosphoric acid (w / w) for 25 to 35 minutes with shaking, rinse with sterile water 5 to 6 times, and shake and soak for 10 to 15 minutes each time; drain, use sterile filter paper to absorb the surface water droplets, and sprinkle them into M6 germination induction medium respectively; place the culture bottle at a temperature of 25°C, a light intensity of 1200Lx, and a photoperiod of 14 hours of light / 10 hours of darkness for 30 days.

[0047] Example 3: Comparison of the germination-promoting effects of metaphosphoric acid and concentrated sulfuric acid on Castanopsis chinensis seeds

[0048] Two-year-old tung oil seeds were used as materials to compare the differences in their germination-promoting effects between metaphosphoric acid and concentrated sulfuric acid. First, the seeds were repeatedly rubbed and rinsed with detergent to remove the wax on the surface of the seeds, and the seeds were shaken and soaked in 100mg / LGA3 solution on a shaker with an oscillation frequency of 200r / min for 24h. After that, they were transferred to a clean bench and soaked in 75% alcohol for 30s, rinsed with sterile water 3 to 5 times, and shaken and soaked for 3 to 5min / time; after draining, the seeds were soaked in sodium hypochlorite with 8% effective chlorine for 30min, rinsed with sterile water 3 to 5 times, and shaken and soaked for 8 to 10min / time; after draining and dividing equally, the seeds were treated with two methods to promote germination: the metaphosphoric acid group was shaken and soaked in 20% metaphosphoric acid (w / w) for 30min, rinsed with sterile water 5 to 6 times, and shaken. The concentrated sulfuric acid group was shaken and soaked for 10 to 15 minutes each time; the concentrated sulfuric acid group was shaken and soaked in 30% concentrated sulfuric acid (v / v) for 20 minutes, rinsed with sterile water 5 to 6 times, and shaken and soaked for 10 to 15 minutes each time; after that, they were drained, the surface water droplets were absorbed with sterile filter paper, and M6 germination induction medium was sprinkled on them respectively; the culture bottles were placed at a temperature of 25°C, a light intensity of 1200Lx, and a photoperiod of 14 hours of light / 10 hours of darkness for 30 days, and the whitening and growth of the seeds were observed every day.

[0049] The comparison of the sprout-promoting effect of this embodiment is as follows: Figure 5 As shown in the germination curve comparison, it can be seen that the metaphosphoric acid treatment (20% metaphosphoric acid soaking for 30 minutes, Figure 5 middle red line) and classic concentrated sulfuric acid treatment (immersion in 30% concentrated sulfuric acid for 20 minutes, Figure 5 The germination-promoting effects of the two treatments (the green broken line in the middle) are similar, as shown by the similar overall germination rate and germination period. Both can make the Castanopsis chinensis seeds germinate explosively around 10-15 days, and the number of germinated individuals rises linearly; thereafter, as the number of individuals to be germinated decreases, the number of newly germinated seeds decreases; and around the 30th day, the germination process is basically completed. During this period, the germination synchronization after metaphosphoric acid treatment is relatively strong, and the seeds reach the peak of germination around 15 days, after which the number of newly germinated seeds decreases smoothly; while after concentrated sulfuric acid treatment, the number of newly germinated seeds will show a small fluctuation increase in the later period, indicating that its germination synchronization is slightly lower than that of the metaphosphoric acid treatment group, and the consistency is slightly worse. It can be seen that metaphosphoric acid treatment can replace concentrated sulfuric acid treatment for Castanopsis chinensis seed germination promotion.

[0050] At the same time, it should be noted that concentrated sulfuric acid is not only a strictly controlled high-risk reagent, that is, it cannot be purchased by registered units that do not have the qualification to purchase hazardous chemicals; and if the used concentrated sulfuric acid waste liquid is accidentally mixed with the sodium hypochlorite waste liquid produced by the previous disinfection during the operation, it will strongly release irritating, corrosive and harmful gases - chlorine gas, which seriously threatens the eyes and respiratory health of nearby people. Strict attention must be paid to safety prevention and control, and special attention must be paid to the classified discharge, storage and recycling of hazardous chemical waste liquids. Relatively speaking, because metaphosphoric acid is a generally controlled reagent and is a stable solid, its procurement, transportation, storage and recycling are more convenient, and its operational safety and simplicity (easy to clean) are significantly higher than concentrated sulfuric acid. It can be seen that metaphosphoric acid treatment provides a more efficient and safer new solution for the breeding of Castanopsis chinensis seedlings.

[0051] The above specific embodiments do not limit the scope of protection of the present invention. Those skilled in the art will appreciate that various modifications, combinations, sub-combinations, and substitutions may occur depending on design requirements and other factors. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention are intended to be included within the scope of protection of the present invention.

Claims

1. A method for inducing germination of Castanopsis chinensis seeds, comprising the following steps: (1) removing wax from the surface of the Castanopsis chinensis seeds, washing them, and then filtering them to obtain wax-removed Castanopsis chinensis seeds; (2) placing the elm-shaped elm-shaped seeds after wax removal in step (1) in a 0-200 mg / L gibberellin GA3 solution, shaking and soaking for 24-36 hours, washing and filtering to obtain GA3-treated elm-shaped elm-shaped seeds; (3) sterilizing the GA3-treated Castanopsis chinensis seeds in step (2), placing them in a metaphosphoric acid aqueous solution with a mass concentration of 10% to 50%, shaking and soaking them for 25 to 45 minutes, performing a germination-promoting treatment, washing them, and filtering them to obtain Castanopsis chinensis seeds after the germination-promoting treatment; (4) Sowing the Castanopsis chinensis seeds after the germination promotion treatment in step (3) into a germination induction medium for germination induction.

2. The method for inducing germination of Castanopsis chinensis seeds according to claim 1, wherein: The mass concentration of the metaphosphoric acid aqueous solution in step (3) is 10% to 30%.

3. The method for inducing germination of Castanopsis sylvestris seeds according to claim 1 or 2, characterized in that: The solution is shaken and soaked in the metaphosphoric acid aqueous solution in step (3) for 25 to 35 minutes.

4. The method for inducing germination of Castanopsis chinensis seeds according to claim 1, wherein: The germination induction medium in step (4) is based on 1 / 2MS, and is supplemented with 0.2 mg / L to 1.0 mg / L of 6-BA, 0.1 mg / L to 0.3 mg / L of IBA, 25 g / L to 30 g / L of sucrose, and 7.0 g / L to 8.0 g / L of carrageenan; the pH value of the germination induction medium is 5.8 to 6.

0.

5. The method for inducing germination of Castanopsis chinensis seeds according to claim 4, characterized in that: The germination induction medium is based on 1 / 2MS, and is supplemented with 1.0 mg / L 6-BA, 0.3 mg / L IBA, 25 g / L sucrose, and 7.0 g / L carrageenan.

6. The method for inducing germination of Castanopsis truncatula seeds according to claim 1 or 4, characterized in that: The germination induction in step (4) is to culture the Castanopsis schrenkiana seeds under conditions of a temperature of 25°C to 28°C, a light intensity of 1200 to 1800 Lx, and a photoperiod of 14 hours of light / 10 hours of darkness for 10 to 60 days.

7. The method for inducing germination of Castanopsis chinensis seeds according to claim 6, characterized in that: The germination induction is to culture the Castanopsis schrenkiana seeds for 30 days under the conditions of a temperature of 25° C., a light intensity of 1200 Lx, and a photoperiod of 14 hours of light / 10 hours of darkness.

8. The method for inducing germination of Castanopsis chinensis seeds according to claim 1, characterized in that: The method for removing the wax on the surface of the Castanopsis chinensis seeds in step (1) is as follows: placing the Castanopsis chinensis seeds in a cloth bag, adding detergent, rubbing and rinsing repeatedly until foam is produced and is not yellow, so as to fully remove the wax on the surface of the Castanopsis chinensis seeds.

9. The method for inducing germination of Castanopsis chinensis seeds according to claim 1, characterized in that: The sterilization method in step (3) is: soaking the Castanopsis schrenkiana seeds treated with gibberellins in 75% alcohol for 30 to 40 seconds, and rinsing them with sterile water for 3 to 4 times; soaking them in a sodium hypochlorite solution with an effective chlorine mass concentration of 6% to 8% for 30 to 40 minutes, and rinsing them with sterile water for 3 to 4 times to completely disinfect the Castanopsis schrenkiana seeds.

10. The method for inducing germination of Castanopsis chinensis seeds according to claim 1, characterized in that: The Castanopsis aviculare seeds are new seeds or old seeds with a storage period of ≤8 years.

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