A method for promoting germination of aloe vera seeds

Abstract

The present application relates to the field of medicinal and edible plant cultivation and breeding, and particularly relates to a method for promoting seed germination of Alocasia macrorrhiza. The present application is characterized in that the Alocasia macrorrhiza seeds are soaked for 24 hours, and then are transferred to a solid culture medium for low-temperature treatment; alternatively, the seeds are directly subjected to low-temperature treatment, and then are transferred to a solid culture medium for germination culture. The method provided by the present application shortens the germination time of the Alocasia macrorrhiza seeds, improves the germination rate of the Alocasia macrorrhiza seeds, and achieves the germination of the Alocasia macrorrhiza seeds in about 35 days, and the germination rate reaches 86.67% within 40 days. The Alocasia macrorrhiza seedlings obtained by the method provided by the present application are sterile seedlings, which can effectively prevent the degeneration of the Alocasia macrorrhiza seeds, reduce the occurrence of diseases and insect pests, and are suitable for large-scale seedling raising. The method provided by the present application is not limited by seasons, effectively solves the problems of low germination rate and long seedling period of the Alocasia macrorrhiza during seed propagation, and provides an effective way for breeding of Alocasia macrorrhiza through sexual reproduction.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of medicinal and edible plant cultivation and breeding, and particularly relates to a method for promoting seed germination of Zingiber mioga Rose. BACKGROUND

[0002] Zingiber mioga Rose, also known as Zingiber mioga Rose, Yanghe, and wild ginger, is a perennial rhizomatous herb of Zingiberaceae, and has the medicinal effects of activating blood and regulating menstruation, relieving cough and reducing phlegm, detoxifying, and invigorating the stomach. According to the Vegetable Volume of China Agricultural Encyclopedia, Zingiber mioga Rose is rich in nutrients such as protein, cellulose, vitamins, and carotene, and its rhizomes, flower buds, and tender buds can be used for medicinal or edible purposes. It is a rare characteristic vegetable with medicinal and edible properties, and is known as the “Asian ginseng”. Zingiber mioga Rose originated in Southeast China and is cultivated in countries such as South Korea, Japan, Australia, and New Zealand. In China, Zingiber mioga Rose is mainly distributed in Yunnan, Guizhou, Chongqing, Anhui, and Jiangsu. In recent years, the demand for Zingiber mioga Rose in the domestic and foreign markets has been increasing, which has prompted the rapid expansion of the area of artificially cultivated Zingiber mioga Rose.

[0003] The main propagation methods of Zingiber mioga Rose include division propagation, seed propagation, and tissue culture, and the division propagation of underground stems is mostly used for seedling breeding in production. However, when the rhizomes are used for asexual propagation, a large amount of germplasm material is required, and it is difficult to achieve large-scale production. In addition, due to the accumulation of diseases and pests, the degeneration of species is easy to occur, which causes the growth potential to weaken, the yield and quality to decrease, and the stress resistance to reduce. Although Zingiber mioga Rose can flower and set seeds, the seed germination rate is low, the growth rate is slow, and the seedling period is long when Zingiber mioga Rose is propagated by seeds under natural conditions. There are few reports on the cultivation method of Zingiber mioga Rose seeds.

[0004] For a long time, the breeding of Zingiber mioga Rose mainly focuses on the selection and utilization of natural variation, and new varieties cannot be effectively selected through sexual hybridization technology, which hinders the process of new variety selection of Zingiber mioga Rose. Therefore, it is of great significance to study the method for promoting the germination of Zingiber mioga Rose seeds for the breeding of excellent varieties of Zingiber mioga Rose and the sustainable and healthy development of the Zingiber mioga Rose industry. SUMMARY

[0005] The present application provides a method for promoting the germination of Zingiber mioga Rose seeds.

[0006] The Zingiber mioga Rose seed is long oval, the seed coat is thick, and there is a thick endosperm between the seed coat and the embryo, which has a dormancy characteristic. Under natural conditions, the seedling period of Zingiber mioga Rose seed propagation is 180-220 days. In nature, the leaching of heavy rain can remove the inhibition and break the dormancy, while in agricultural production, the methods such as flushing, rinsing and soaking, cold treatment, peeling the embryo (removing the seed coat), and treatment with GA3 hormone are mainly used to promote germination.

[0007] The present application carries out a large number of experimental work to promote the germination of Alocasia seed. The most suitable method for Alocasia seed germination is accidentally discovered.

[0008] For seeds with thick seed coat, the prior art usually adopts the method of seed soaking. However, for Alocasia seed, water or gibberellin seed soaking for 24 hours and then sowing in the seedling substrate, no germination is observed within 60 days. The inventor refers to in vitro vegetative propagation, and replaces the seedling substrate with solid culture medium. However, water or gibberellin seed soaking for 24 hours and then inoculating into the solid culture medium, it is found that Alocasia seed still has no germination within 60 days.

[0009] The inventor adjusts the experiment again. On the basis of water or gibberellin seed soaking for 24 hours, other treatment methods for breaking seed dormancy are added. The results show that: after Alocasia seed soaking in water for 24 hours, low-temperature treatment is carried out, and after disinfection treatment, it is transferred to the solid culture medium, it can germinate in about 50 days, and the germination rate obtained by statistics on the 50th day is 50%. After Alocasia seed soaking in gibberellin for 24 hours, low-temperature treatment is carried out, and after disinfection treatment, it is transferred to the solid culture medium, it can also germinate in about 50 days, and the germination rate obtained by statistics on the 50th day is 46.67%.

[0010] The present application first discovers that for Alocasia seed, after seed soaking, the order of seed low-temperature treatment and seed inoculation into the solid culture medium will have a great influence on the time required for seed germination and the germination rate.

[0011] For example, after water soaking, the seed is inoculated into the culture medium first, and the seed and the culture medium are subjected to low-temperature treatment together, which can improve the germination rate of Alocasia seed, and the germination rate obtained by statistics on the 50th day is 76.67%, which is higher than 50%. After gibberellin soaking, the seed is inoculated into the culture medium first, and the seed and the culture medium are subjected to low-temperature treatment together, which can shorten the germination time to about 40 days.

[0012] Therefore, under the condition of seed soaking, after the seed and the solid culture medium are subjected to low-temperature treatment together, the germination effect of Alocasia seed can be significantly improved. After seed soaking, the germination state of Alocasia seed is relatively consistent, which is conducive to batch processing and operation of Alocasia sterile seedlings. At the same time, for the seed with surface damage, after seed soaking and low-temperature treatment, the germination potential of the seed can be effectively maintained.

[0013] The present application also accidentally discovers that without seed soaking, after Alocasia seed is disinfected and inoculated into the solid culture medium, and then subjected to low-temperature treatment, the germination time can also be shortened to about 40 days, and the germination rate obtained by statistics on the 40th day is 46.67%, which is higher than the germination rate in the case of gibberellin seed soaking.

[0014] It is further found that, after omitting seed soaking, the order of low-temperature treatment and inoculation into solid culture medium needs to be adjusted to achieve better germination of the Alocasia seed.

[0015] Therefore, the application provides a method for promoting germination of Alocasia seed, which comprises the following steps:

[0016] Alternatively, seed soaking is omitted, the Alocasia seed is first subjected to low-temperature treatment, and then is transferred into solid culture medium for germination culture after sterilization.

[0017] In the method, the pH of the solid culture medium is 5.8-6.0, and the solid culture medium is obtained by adding 25-30 g / L sucrose and 7-9 g / L agar to MS basic medium.

[0018] It is found that the solid culture medium obtained by adding 25-30 g / L sucrose and 7-9 g / L agar to MS basic medium is the most suitable substrate for germination of the Alocasia seed.

[0019] In the method, the low-temperature treatment is cold storage at 3-5 DEG C for 5-7 days.

[0020] In the method, the sterilization is performed by using a sodium hypochlorite solution with a volume concentration of 4.0-6.5% for 5-10 min.

[0021] In the method, the sterilization is performed by using a sodium hypochlorite solution with a volume concentration of 4.0-6.5% for 5-10 min.

[0022] In the method, the germination culture condition is temperature of 25-30 DEG C and illumination of 1500-2000 lx / 12 h.

[0023] The method for promoting germination of Alocasia seed comprises the following steps:

[0024] (1) selecting normal and fully mature Alocasia seed grains, removing abnormal seed grains and impurities, and airing the seed grains for 2-3 days in advance;

[0025] Airing the seed grains can kill the bacteria attached to the surface of the seed grains by using short-wave light in the sunlight spectrum, and can also improve the activity of enzymes in the seed grains, thereby improving the germination potential and making the seed grains sprout fast and uniformly.

[0026] Most plants are with starch or fat as energy storage material, to provide nutrition for the initial germination of plants. The same is true for plants, only energy storage material is not enough for it to germinate, only mixed with air can convert energy storage material into the required energy to make it germinate.

[0027] (2) omitting seed soaking, the seeds are refrigerated at 3-5℃ for 5-7 days;

[0028] (3) removing the white perisperm on the edge of the seed and the attachments on the surface of the hilum, and rinsing with running water for 4-6 hours after cold treatment;

[0029] In the present application, the white perisperm on the edge of the seed and the attachments on the surface of the hilum are removed after low-temperature treatment, which can protect the seeds during low-temperature treatment and further remove the obstacles to germination.

[0030] (4) rinsing the seeds with sterile water for 3-5 times after disinfection treatment with 75% ethanol for 1 min;

[0031] (5) rinsing the seeds with sterile water for 3-5 times after disinfection treatment with 4.0-6.5% sodium hypochlorite solution for 8-10 min;

[0032] (6) drying the water on the surface of the seeds, and inoculating the seeds on the sterilized solid medium, and culturing the seeds under the conditions of temperature 25-30℃ and light 1500-2000lx / 12h.

[0033] Alternatively, the method for promoting the germination of the seed of the plantain provided by the present application comprises:

[0034] (1) selecting normal development and fully matured plantain seed grains, removing abnormal seed grains and impurities, and airing for 2-3 days in advance;

[0035] (2) soaking the seeds in water or gibberellin for 24h;

[0036] (3) removing the white perisperm on the edge of the seed and the attachments on the surface of the hilum, and rinsing with running water for 4-6 hours;

[0037] (4) disinfecting the seeds with 75% ethanol for 1 min, and rinsing with sterile water for 3-5 times;

[0038] (5) rinsing the seeds with sterile water for 3-5 times after disinfection treatment with 4.0% sodium hypochlorite solution for 8 min;

[0039] (6) drying the water on the surface of the seeds, and inoculating the seeds on the sterilized solid medium;

[0040] (7) refrigerating the inoculated medium at 3-5℃ for 5-7 days;

[0041] (8) The seed germination is carried out at a temperature of 25-30 DEG C and under illumination of 1500-2000 lx / 12 h.

[0042] In the second aspect, the application provides a cultivation method of Agave americana, wherein the seedlings of Agave americana are obtained by the above method, and the seedlings are transplanted when the roots of the aseptic seedlings are 2-3 cm long, the number of roots is 6-10, the height of the seedlings is 5-6 cm, and 3-4 leaves are fully expanded.

[0043] The aseptic seedlings obtained by the method are transplanted into a seedling tray filled with a substrate of peat soil: perlite (5:1) after hardening, and then sprayed with a 50% wettable powder carbendazim solution at a concentration of 0.001%, and covered with a plastic film to maintain a humidity of more than 80%. After 25 days, the survival rate can reach 90%. The virus-free seedlings in the seedling tray are planted in the field when the seedlings have 6-8 leaves and new roots grow fully. When the seedlings are transplanted in the field, the seedlings are watered thoroughly to fix the roots, and the seedlings are shaded (the shading degree is 50%). The seedlings are managed according to the daily management of Agave americana.

[0044] In the third aspect, the application also provides the use of the above method or the above cultivation method of Agave americana in promoting the sexual reproduction of Agave americana.

[0045] The application has the following beneficial effects:

[0046] The application provides two methods for improving the seed germination rate of Agave americana, i.e., under the condition of seed soaking, the seeds are subjected to low-temperature treatment together with a solid culture medium, so that the seed germination effect of Agave americana is significantly improved.

[0047] Under the condition of omitting seed soaking, the seeds are inoculated into the solid culture medium after cold storage, so that the germination time of the seeds is shortened to about 35 days, the germination rate reaches 86.67% within 40 days, and the germination uniformity is high, which is suitable for large-scale seedling raising.

[0048] The seedlings of Agave americana obtained by the method are aseptic seedlings, which can effectively prevent the degeneration of the seed nature of Agave americana and reduce the occurrence of diseases and pests.

[0049] The method provided by the application is simple and efficient, and the operation is not limited by seasons. The method can effectively solve the problems of low seed germination rate and long seedling period of Agave americana, and provides an effective way for breeding improved varieties of Agave americana by sexual reproduction, and has good economic and social benefits. DETAILED DESCRIPTION

[0050] In order to make the objects, technical solutions and advantages of the present application clearer, the technical solutions in the present application will be clearly and completely described below. Obviously, the described embodiments are some of the embodiments of the present application, but not all the embodiments. Based on the embodiments in the present application, all other embodiments obtained by those skilled in the art without creative work fall within the protection scope of the present application.

[0051] The solid culture medium used in the present application is obtained by adding 30 g / L sucrose and 7-9 g / L agar to MS basic medium.

[0052] The MS medium, sodium hypochlorite solution and seedling raising substrate used in the present application are purchased, and the main raw materials of the seedling raising substrate include peat, cotton seed hulls, coconut shells, carbonized rice hulls, sugar cane residues and wood chips.

[0053] The solvent used for seed soaking in the present application is water or gibberellin.

[0054] Embodiment 1

[0055] This embodiment provides the influence of different treatment methods on the germination of Ensete glaucum seeds, and each treatment group in this embodiment includes three repetitions. Treatment groups 1 and 2 are seed germination treatments without seed soaking.

[0056] In the treatment group 1, the steps are as follows:

[0057] (1) Select normally developed and fully matured Ensete glaucum seed grains, remove alien seed grains and impurities, and dry for 2-3 days in advance;

[0058] (2) Skip seed soaking;

[0059] (3) Store the seeds at 4℃ for 7 days;

[0060] (4) After the cold storage treatment, remove the white false seed coat on the edge of the seeds and the attachments on the surface of the seed hilum, and rinse with running water for 4-6 hours;

[0061] (5) Sterilize the seeds with 75% ethanol for 1 min, and wash with sterile water for 3 times;

[0062] (6) Sterilize the seeds washed with sterile water with 4.0% sodium hypochlorite solution for 8 min, and wash with sterile water for 3 times;

[0063] (7) Absorb the water on the surface of the seeds, and inoculate the seeds into the sterilized solid culture medium;

[0064] (8) Perform germination culture under the conditions of temperature 25-30℃ and light intensity 1500-2000 lx / 12 h, and obtain sterile seedlings.

[0065] (9) When the root length of the aseptic seedling is 2-3 cm, the root number is 8-10, the plant height is 5-6 cm, and 3-4 fully expanded leaves, the seedling is hardened and transplanted.

[0066] The steps of the treatment group 2 are as follows:

[0067] (1) Select normal development and fully matured seed grains of the seed, remove alien seed grains and impurities, and air dry for 2-3 days in advance;

[0068] (2) Skip seed soaking;

[0069] (3) Remove the white perisperm on the edge of the seed and the appendages on the surface of the seed navel, and rinse with running water for 4-6 hours;

[0070] (4) Sterilize the seed with 75% ethanol for 1 min, and wash with sterile water for 3 times;

[0071] (5) Sterilize the seed washed with sterile water with 4.0% sodium hypochlorite solution for 8 min, and wash with sterile water for 3 times;

[0072] (6) Dry the water on the surface of the seed, and inoculate the seed on the sterilized solid culture medium;

[0073] (7) Store the inoculated culture medium at 4°C for 7 days;

[0074] (8) Perform germination culture under the condition of temperature 25-30°C and illumination 1500-2000 lx / 12 h to obtain the aseptic seedling.

[0075] (9) When the root length of the aseptic seedling is 2-3 cm, the root number is 8-10, the plant height is 5-6 cm, and 3-4 fully expanded leaves, the seedling is hardened and transplanted.

[0076] Treatment groups 3-6 are seed soaking with water or gibberellin and low-temperature treatment for seed germination treatment.

[0077] The purpose of seed soaking is to make the seed quickly absorb water to reach the water content for normal germination. The water content of dry seed is usually below 15%, and the physiological activity is very weak, in a dormant state. After the seed absorbs water, the seed coat swells and softens, the oxygen dissolved in water enters the cells with water, and the enzymes in the seed also begin to activate. Due to the action of enzymes, the respiration of the embryo is enhanced, and the insoluble substances stored in the endosperm are gradually converted into soluble substances and transported to the embryo with water. The embryo obtains water, energy and nutrients, and under suitable temperature and oxygen conditions, the cells begin to divide, elongate and break through the seed coat (germination).

[0078] The difference between the treatment group 3 and the treatment group 2 is that the step (2) of the treatment group 3 is: water-soaking the seed of the seed for 24 h.

[0079] The difference between the treatment group 4 and the treatment group 2 is that the step (2) of the treatment group 4 is that the gibberellin is used to soak the seeds of the Alocasia macrorrhiza for 24 hours, and the concentration of the gibberellin is 100 mg / L.

[0080] The difference between the treatment group 5 and the treatment group 1 is that the step (2) of the treatment group 5 is that the seeds of the Alocasia macrorrhiza are soaked in water for 24 hours.

[0081] The difference between the treatment group 6 and the treatment group 1 is that the step (2) of the treatment group 6 is that the gibberellin is used to soak the seeds of the Alocasia macrorrhiza for 24 hours, and the concentration of the gibberellin is 100 mg / L.

[0082] In the embodiment, the seed germination effects of the treatment groups are shown in Table 1.

[0083] Table 1 Seed germination effects of Alocasia macrorrhiza

[0084]

[0085]

[0086] From the results of Table 1, it can be seen that:

[0087] (1) The combination of water soaking and low-temperature treatment can shorten the germination time to about 50 days, but the 50-day germination period is relatively high in terms of time cost for breeding. The inventors continue to explore the seed germination of Alocasia macrorrhiza and find that the combination of gibberellin soaking and low-temperature treatment in the treatment group 4 can further shorten the germination time to about 40 days.

[0088] (2) From the results of the treatment groups 1-2, even if the soaking is omitted, the order of low-temperature treatment and medium culture needs to be adjusted. If the soaking experience is followed, the seeds are first transferred to the medium and then subjected to low-temperature treatment, the Alocasia macrorrhiza seeds cannot be well germinated.

[0089] (3) From the comparison of the treatment groups 3-4 and 5-6, it can be seen that after the seeds of Alocasia macrorrhiza are soaked, the seeds are subjected to low-temperature treatment together with the solid medium, and the seed germination effect is better than that of the seeds subjected to low-temperature treatment alone and then inoculated into the medium.

[0090] Germination effect of the seeds of the Alocasia macrorrhiza using the seedling substrate in the comparative example 1

[0091] In the comparative example, each treatment group includes three repetitions. In the comparative example, the treatment conditions and seed germination effects of the treatment groups are shown in Table 2.

[0092] Table 2 Seed germination effects of Alocasia macrorrhiza

[0093]

[0094]

[0095] The present comparative example investigates different methods of breaking seed dormancy, normal temperature seed soaking, gibberellin treatment, low temperature treatment, etc. The results in Table 2 show that no matter what treatment is used to break dormancy, the Alocasia seed is not germinated within 60 days when sown in the seedling substrate. The inventors believe that the conventional sexual reproduction seedling substrate is not suitable for the germination of Alocasia seeds.

[0096] Example 2 Alocasia seed germination using solid medium

[0097] The inventors, in combination with the biological characteristics of Alocasia seeds, prepared a solid medium by adding 25-30 g / L sucrose and 7-9 g / L agar to the MS basic medium, which was used as the substrate for Alocasia seedling culture. Each treatment group in the present comparative example includes three replicates, and the seed germination effect is shown in Table 3.

[0098] Table 3 Alocasia seed germination effect

[0099]

[0100] For seeds with thick seed coats, the prior art usually uses the method of seed soaking. However, the results in Table 3 further confirm that for Alocasia seeds, when using solid medium as the seed germination substrate, omitting seed soaking and using medium culture can achieve better Alocasia seed germination effect, shorter germination time, and higher germination rate than seed soaking treatment.

[0101] The possible reason is that Alocasia seeds have a thick endosperm, and during normal temperature seed soaking, the water used for seed soaking can cause the fat layer to swell, squeezing the growth space of the embryo, and affecting the development of the embryo.

[0102] Example 3 Different sodium hypochlorite solution treatment times

[0103] Each treatment group in the present comparative example includes three replicates, and 20 seeds are used for each replicate.

[0104] Table 4 Alocasia seed medium contamination rate after inoculation

[0105]

[0106] The results in Table 4 show that 4.0% sodium hypochlorite solution treatment for 8 min can achieve a significant sterilization effect compared to treatment for 5 min, and there is no significant difference in sterilization effect compared to treatment for 10 min, and the damage to the seeds is relatively small. Therefore, for Alocasia seeds, the most suitable sterilization method is 4.0% sodium hypochlorite solution treatment for 8 min.

[0107] It should be pointed out finally that the above embodiments are only used to illustrate the technical solutions of the present application, but not to limit the same; and although the present application has been described in detail with reference to the foregoing embodiments, it should be appreciated by those skilled in the art that the technical solutions recorded in the foregoing embodiments can be modified, or some technical features thereof can be replaced equivalently; and these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the spirit and scope of the technical solutions of the embodiments of the present application.

Claims

1. A method of promoting germination of a cattail seed, the method comprising, The method comprises the following steps: (1) selecting normal and mature seeds of Zizania aquatica, removing abnormal seeds and impurities, and airing for 2-3 days in advance; (2) omitting seed soaking, and cold storing the seeds at 3-5 DEG C for 5-7 days; (3) removing the white false seed coat on the edge of the seeds and the attachments on the surface of the seed hilum after the cold storage treatment, and washing the seeds with flowing water for 4-6 hours; (4) sterilizing the washed seeds with 75% ethanol for 1 minute, and washing the seeds with sterile water for 3-5 times; (5) sterilizing the washed seeds with 4.0-6.5% sodium hypochlorite solution for 8-10 minutes, and washing the seeds with sterile water for 3-5 times; (6) drying the water on the surface of the seeds, and inoculating the seeds on sterilized solid culture medium, and culturing the seeds at a temperature of 25-30 DEG C and under illumination of 1500-2000 lx / 12 hours.

2. The application of the method of claim 1 in promoting the sexual reproduction of Zizania aquatica.

3. The application of the method of claim 1 in the breeding of Zizania aquatica.

4. The application of the method of claim 1 in the batch cultivation of Zizania aquatica aseptic seedlings.

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