Prothallus proliferation culture medium and rapid propagation method for rare and endangered plant cibotium barometz

Abstract

The invention discloses a prothallus proliferation culture medium and a rapid propagation method for rare and endangered plant cibotium barometz. The prothallus proliferation culture medium of cibotium barometz is used for accelerating the proliferation of gametophytes, and the prothallus maturation time is shortened; meanwhile, the invention provides a rapid propagation method of cibotium barometz, which comprises the following steps: carrying out artificial sterile culture on cibotium barometz spores, rapidly germinating in a culture bottle to form prothalli, culturing on a prothalli proliferation culture medium, inducing and transforming gametophytes in a specific matrix outside the bottle to form sporophytes by virtue of an incomplete tissue culture method, and transplanting. According to the method, the formation of prothallium is accelerated, the sporophyte conversion rate is increased, the reproduction rate is increased while the domestication and reproduction of sporophyte seedlings are accelerated, a large number of cibotium barometz seedlings which grow healthily can be obtained, the survival rate reaches 90%, the spore breeding period is obviously shortened, the operation requirement is not strict, and the method is suitable for large-scale popularization and application. The technical problems that the seedlings in the tissue culture bottle are easy to pollute, fragile and low in seedling rate are solved.

Description

Technical Field

[0001] The invention belongs to the technical field of biological breeding, and more specifically, relates to a prothallus proliferation culture medium and a rapid propagation method of a rare and endangered plant Cibotium chinense. Background Art

[0002] Cibotium barometz is a traditional folk medicine with a wide range of pharmacological effects, including hemostasis, antibacterial, osteoporosis prevention and treatment, anti-cancer, and liver protection. Cibotium barometz is mainly used as medicine with its rhizome. Due to the large market demand, people over-exploit it, and the limited reserves of wild resources of Cibotium barometz, Cibotium barometz is also listed as a second-level key protected plant by the state. Due to the relatively backward artificial breeding technology and cultivation management methods, the resources of Cibotium barometz have been sharply reduced year by year, and artificial large-scale planting is urgently needed to meet people's needs.

[0003] At present, people usually use the method of ramet propagation to artificially propagate Cibotium chinense, but it is limited by the number of original mother plants, the ramet propagation coefficient is low, and the ramet propagation number is small, the speed is slow, and a large number of seedlings cannot be obtained in a short period of time. Tissue culture is a technology that plant explants are cultivated under artificial control conditions to obtain a complete plant for regeneration. Although the spores are artificially given the most suitable growth conditions, due to the particularity of the material, there are still problems that the gametophyte is difficult to transform into the sporophyte and the conversion rate is low. Sporangium is the reproductive organ of ferns, which contains many reproductive cells (spores). Ripe spores can grow to form gametophytes, and finally form new complete individuals. It is easy to obtain in large numbers and does not cause damage to plants. People also use its spores to carry out tissue culture and propagation. But there is still a problem of spore seedling rate in the natural environment, which can not meet the needs of market and social development.

[0004] The natural propagation of Cibotium chinense is mainly through spore propagation. For example, the prior art discloses a tissue culture method for Cibotium chinense spores, which mainly accelerates the development of prothallus to form sporophytes by optimizing different culture media and plant hormone combinations. However, in this method, the prothallus takes a long time to form sporophytes in the tissue culture bottle, the conversion rate of sporophytes is low, and it is very inconvenient to operate in the bottle. The transformed sporophytes are easily affected by the natural environment such as soil, temperature, light, and population in the original habitat after seedling training and transplanting, which will lead to a decrease in the breeding rate. In addition, in the process of proliferation from prothallus differentiation after spore germination, the normal alternation of generations is affected by plant hormones, salt ion components or culture conditions, etc., and the edges of the prothallus will thicken during the growth of the prothallus, inducing small spherical embryoids, causing the prothallus to proliferate in large quantities to form a prothallus mass, and ultimately unable to form sporophytes. It may also cause incomplete development of the gametophyte, or directly generate sporophytes from the gametophyte. Therefore, in order to study more rapid propagation methods and culture media suitable for the endangered plant Cibotium barometz, accelerate its formation of spore seedlings, shorten the cultivation period, and improve the rapid propagation method of Cibotium barometz, a theoretical basis can be provided for the later large-scale production. Summary of the invention

[0005] The technical problem to be solved by the invention is to overcome the defects and shortcomings in the rapid propagation of Cibotium chinense spore tissue culture, and to provide a prothallus proliferation culture medium and a rapid propagation method of Cibotium chinense, a rare and endangered plant.

[0006] The invention aims to provide a prothallus proliferation culture medium of Cibotium chinense.

[0007] Another object of the present invention is to provide an application of a culture medium for the proliferation of prothallus of Cibotium caninum.

[0008] Another object of the present invention is to provide a method for rapid propagation of Cibotium barometz.

[0009] The above-mentioned purpose of the present invention is achieved through the following technical solutions:

[0010] The invention provides a protothallus proliferation culture medium of Cibotium chinense, which takes 1 / 2MS culture medium as a basic culture medium and further contains the following components: 30g / L sucrose+6g / L agar+50-200g / L natural organic additive+0.1g / L activated carbon, wherein the natural organic additive is banana juice, potato juice or coconut juice.

[0011] The invention uses Cibotium caninum spore powder as a basic material, adds natural organic additives into a culture medium, and adopts a specific proliferation culture medium, so that the prothallus can be obviously proliferated, and the proliferation multiples of the prothallus are significantly increased by 6.69 times. The cultured prothallus mass is emerald green in color and large in volume, and the maturation time of the prothallus is accelerated; the proliferation multiples of the prothallus are increased, which is beneficial to the subsequent transformation of the sporophyte, and is further used in the subsequent incomplete tissue culture method.

[0012] Furthermore, the composition of 1 / 2MS medium is: macroelements KNO 3 950mg / L, NH 4 NO 3 825mg / L, MgSO 4 90.35mg / L, KH 2 PO 4 85mg / L; trace element MnSO 4 ·H 2 O 16.9mg / L, ZnSO 4 7H 2 O 8.6mg / L, H 3 BO 3 6.2mg / L, KI 0.83mg / L, Na 2 MoO 4 ·2H 2 O 0.25mg / L, CoCl 2 6H 2O 0.025mg / L, CuSO 4 ·5H 2 O0.025mg / L; Iron salt FeNaEDTA 36.7mg / L; Organic matter calcium salt CaCl 2 166.1mg / L; Glycine 2.0mg / L; Thiamine hydrochloride 0.1mg / L; Pyridoxine hydrochloride 0.5mg / L; Nicotinic acid 0.5mg / L; Inositol 100mg / L.

[0013] Preferably, the basal culture medium is 1 / 2MS culture medium and further contains the following components: 30 g / L sucrose + 6 g / L agar + 100 g / L natural organic additives + 0.1 g / L activated carbon.

[0014] More preferably, the natural organic additive is banana juice.

[0015] The invention provides application of the proliferation culture medium in rapid propagation of Cibotium caninum.

[0016] The invention uses artificial aseptic culture of Cibotium spores and a prothallus proliferation culture medium of Cibotium spores to quickly germinate and form Cibotium spores in a culture bottle, and uses a specific prothallus proliferation culture medium containing natural organic additives to accelerate the growth and development of the prothallus (gametophyte). Through an incomplete tissue culture method, the gametophyte is induced to transform into a sporophyte in an optimal matrix outside the bottle, the spore breeding cycle is shortened, the sporophyte seedling domestication is accelerated, a large number of healthy Cibotium spores are obtained, technical support is provided for artificial large-scale planting of Cibotium spores, reference is provided for the sustainable development of Cibotium spore medicinal material planting and application, and Cibotium spores plants are artificially planted on a large scale.

[0017] Therefore, the present invention provides a method for rapid propagation of Cibotium chinense, wherein Cibotium chinense spores are germinated and cultured, and then inoculated into a prothallus proliferation culture medium of Cibotium chinense for proliferation culture, and the prothallus after proliferation is transferred to a specific matrix for indoor culture, and then transplanted. The specific method is as follows:

[0018] S1. Take the spore powder of Cibotium caninum, disinfect it, suspend it with sterile water, and then inoculate it in a solid culture medium for germination culture;

[0019] S2. The young prothallus clusters with a growth time of more than 30 days in S1 are divided into small prothallus clusters, inoculated into the prothallus proliferation medium of Cibotium serrata, and rapidly proliferated to obtain the prothallus clusters after proliferation;

[0020] S3. After the proliferated prothallus mass is sheared, it is transferred to a sterilized peat soil-vermiculite matrix for further cultivation to obtain a sporophyte seedling;

[0021] S4. Transplant the sporophyte seedlings into a peat soil-vermiculite-perlite matrix, culture them to keep them moist, and then culture them in the usual way after two weeks to obtain young sporophyte seedlings.

[0022] Preferably, in step S1, the solid culture medium is selected from 1 / 2MS solid, 1 / 4MS solid or 1 / 4MS solid + 1% sucrose; the culture conditions are: fluorescent light source, temperature 25-27° C., and illumination time 12 h / d.

[0023] More preferably, the solid culture is 1 / 4MS solid.

[0024] Preferably, the method for disinfecting the spore powder and treating the sterile water suspension in step S1 is as follows: taking the spore powder, soaking it in 75% alcohol for 60 seconds for disinfection, and then centrifuging to remove the supernatant; then soaking it in 4% NaClO for 20 minutes, and then centrifuging to remove the supernatant; finally, washing it with sterile water for 4 to 6 times, and centrifuging to remove the supernatant, and finally preparing 1.5 mL of spore suspension.

[0025] Further preferably, the proliferation medium in step S2 contains 30 g / L sucrose + 6 g / L agar + 100 g / L banana juice + 0.1 g / L activated carbon.

[0026] Preferably, the shearing treatment in step S3 is: mixing the protothallus mass with 1 / 2MS salt solution, stirring and shearing for 5 seconds; and the dosage ratio of the peat soil-vermiculite matrix is ​​1:1.

[0027] Preferably, the moisturizing culture in step S3 is: spraying Japanese garden formula nutrient solution every other day; the formula of the Japanese garden formula nutrient solution is: 945 mg / L calcium nitrate tetrahydrate + 809 mg / L potassium nitrate + 153 mg / L diammonium hydrogen phosphate + 493 mg / L magnesium sulfate heptahydrate.

[0028] Preferably, in step S4, the sporophyte seedlings are 3 to 5 cm in height and have 2 to 3 true leaves.

[0029] The rapid propagation method of Cibotium barometz provided by the present invention is different from the key technology of the maturation of gametophytes and the transformation process to sporophytes in the traditional tissue culture method. The traditional tissue culture method accelerates the development of prothallus to form sporophytes, while the present invention uses spore powder as a basic material, accelerates its growth and development by tissue culture (using a specific prothallus proliferation culture medium), improves the transformation rate of sporophytes by an incomplete tissue culture method, shortens the time for seedling hardening and acclimatization, accelerates its reproduction and improves the reproduction rate at the same time, and has no strict operation requirements, thereby solving the technical problems that seedlings are easily contaminated, fragile, and have a low seedling rate in tissue culture bottles.

[0030] The present invention has the following beneficial effects:

[0031] The invention provides a prothallus proliferation culture medium of a rare and endangered plant Cibotium chinense. Cibotium chinense spore powder is used as a basic material. By adding natural organic additives to the culture medium, a specific proliferation culture medium can be used to significantly proliferate the prothallus, accelerate the prothallus proliferation speed, and significantly increase the prothallus proliferation multiple by 6.69 times. The cultured prothallus mass is emerald green in color and large in volume, which accelerates the prothallus maturation time. The prothallus proliferation culture medium of Cibotium chinense provided by the invention can significantly increase the prothallus proliferation multiple, is conducive to the subsequent transformation of sporophytes, and is used in the rapid propagation of Cibotium chinense spore tissue culture, can shorten the prothallus proliferation time, and further accelerate the formation of sporophytes in the tissue culture bottle.

[0032] At the same time, the present invention provides a method for rapid propagation of the rare and endangered plant Cibotium chinense. The method comprises the following steps: artificial aseptic culture of Cibotium chinense spores, rapid germination of Cibotium chinense spore powder in a culture bottle, culture in a prothallus proliferation culture medium of Cibotium chinense, and accelerating the growth and development of its gametophyte (prothallus) by using natural organic additives; and in an incomplete tissue culture method, inducing the gametophyte to transform into a sporophyte in a specific matrix outside the bottle, cultivating sporophyte seedlings, and transplanting the young sporophyte seedlings to grow healthily. The present invention cultivates the prothallus mass through a specific culture matrix, can significantly shorten the transformation time of the sporophyte, improve the sporophyte transformation rate, and further shorten the time for seedling hardening and taming, accelerates its reproduction and improves the reproduction rate at the same time, and can obtain a large number of healthy Cibotium chinense seedlings, the survival rate of which reaches 90%. In addition, the method provided by the present invention is not very strict in terms of operation requirements, can transform the sporophyte in the matrix after the prothallus proliferation culture, and solves the technical problems that the seedlings are easily contaminated, relatively fragile, and have a low seedling rate in the tissue culture bottle.

[0033] The invention provides technical support for the large-scale artificial planting of Cibotium barometz and provides a reference for the sustainable development of the planting and application of Cibotium barometz medicinal materials. Large-scale artificial planting of Cibotium barometz plants. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Figure 1 It is mature spore material scattered naturally;

[0035] Figure 2 The effects of different natural additives on the proliferation of Cibotium cichorium protothallus (A: banana juice; B: potato juice; C: coconut juice);

[0036] Figure 3 The transformation effects of different substrates inducing sporophytes (A: river sand-perlite; B: peat soil-vermiculite; C: peat soil-perlite; D: peat soil-river sand-perlite);

[0037] Figure 4The transformation effects of different substrates on sporophytes after 60 days of induction (A: river sand-perlite; B: peat soil-vermiculite; C: peat soil-perlite; D: peat soil-river sand-perlite);

[0038] Figure 5 The diagram shows the growth of a single sporophyte after transplantation (A: young sporophyte seedlings after transplantation; B: young sporophyte seedlings that have grown for a period of time; C: a single sporophyte growing in a substrate). DETAILED DESCRIPTION

[0039] The present invention is further described below in conjunction with the accompanying drawings and specific examples, but the examples do not limit the present invention in any form. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the art.

[0040] Unless otherwise specified, the reagents and materials used in the following examples are commercially available.

[0041] Sucrose-free 1 / 4MS solid, 1 / 2MS solid, MS solid, and MS liquid media were purchased from Beijing Solebow Technology Co., Ltd.;

[0042] The composition of the MS medium (containing no agar and sucrose) used in the present invention is shown in the following table:

[0043] Table 1 MS culture medium composition

[0044]

[0045] Table 2 1 / 2MS culture medium composition

[0046]

[0047]

[0048] Table 3 1 / 4MS culture medium composition

[0049]

[0050] The present invention collects mature spore leaves of sporangium and collects naturally scattered mature spores as materials, which are collected in Dinghushan Nature Reserve, Zhaoqing, Guangdong. They are obtained through legal collection and used for the following research (with a collection certificate issued locally).

[0051] Example 1 Effect of Inorganic Salt Concentration on Spore Germination of Cibotium chinense

[0052] About 10 mg of Cibotium spores ( Figure 1) After being soaked and disinfected with 75% alcohol for 60 seconds, the supernatant was removed by centrifugation. The precipitate was soaked and disinfected with 4% sodium hypochlorite for 20 minutes and then centrifuged. The precipitate was washed with sterile water for 4 to 6 times, and finally sterile water was added to make a 1.5 mL suspension, which was inoculated into 1 / 4 MS solid, 1 / 2 MS solid, MS solid and MS liquid culture media without sucrose. 0.5 mL was inoculated into each bottle, 15 bottles were inoculated for each type, and the process was repeated 3 times. The spores were cultured in a culture room under a fluorescent lamp for 12 hours and in darkness for 12 hours. The appearance of green dots on the surface of the culture medium indicated spore germination, and the earliest spore germination time and germination rate were recorded.

[0053] The results are shown in Table 4, which show that the spores of Cibotium chinense can germinate normally in different basic culture media, and different inorganic salt concentrations have significant differences in spore germination time. Among them, the spores in 1 / 4MS solid culture medium germinate the earliest, the shortest is 12 days, the germination rate reaches 100%, and the germination time and end time are significantly shorter than those in MS liquid culture medium. Cibotium chinense spores are suitable for germination in low concentrations of inorganic salts, and sucrose has no significant effect on spore germination, but the appropriate culture medium can shorten the germination culture time.

[0054] Table 4 Effects of different culture media on spore germination of Cibotium caninum (n=3)

[0055]

[0056] Note: Different lowercase letters in the same column indicate significant differences (P<0.05).

[0057] Example 2 Effects of different natural additives on the proliferation of Cibotium chinense protothallus

[0058] The young prothallus clusters formed after germination for 60 days were carefully removed with sterilized tweezers, divided into small pieces with a diameter of 1 cm (about 0.15 g), and inoculated into proliferation culture media containing different amounts of natural organic additives: 1 / 2MS + 30 g / L sucrose + 6 g / L agar + different natural organic additives + 0.1 g / L activated carbon, wherein the different natural organic additives are banana juice, potato juice, and coconut juice (the raw materials can be purchased commercially, wherein the banana juice and potato juice are obtained by chopping potatoes and bananas, boiling them, and removing the filter residue) and set different amounts: 50, 100, 150, and 200 g / L; 4 groups (about 0.6 g) were inoculated per dish, repeated 3 times, and the initial total weight of the prothallus groups was recorded. After 30 days of culture, the prothallus groups were taken out and weighed, and the total weight of the prothallus groups after proliferation was recorded, and the proliferation multiple = (total weight of prothallus after proliferation - total weight of prothallus before proliferation) / total weight of prothallus before proliferation.

[0059] The results are shown in Table 5 below. The prothallus in the culture medium with different natural organic additives showed significant proliferation ( Figure 2), among which the proliferation multiple of banana juice with 100g / L was the highest, reaching 6.69 times, the prothallus was dark green and larger in size, which accelerated the maturation time of the prothallus. The prothallus could proliferate in the three natural additives, and the proliferation multiples increased first and then decreased with the concentration of additives, reaching the maximum value in the concentration of 100g / L, the prothallus group was emerald green in color and larger in size.

[0060] Table 5 Effects of different natural additives on the proliferation of the protothallus of Cibotium chinense (n=3)

[0061]

[0062] Note: The data in the table are mean ± standard deviation. Different lowercase letters in the same column indicate significant differences (P < 0.05).

[0063] Therefore, 100 g / L banana was selected as a natural organic additive for the subsequent incomplete tissue culture method to shorten the time of seedling hardening and acclimatization, accelerate its reproduction and improve its reproduction rate.

[0064] Example 3 Effects of different substrates on the transformation of Cibotium chinense sporophytes

[0065] The protothallus mass after proliferation in Example 2 was transferred to a fresh 1 / 2MS basal medium without organic additives, and continued to adapt and culture for one week. After that, the cover was opened and adapted for 3 to 5 days, the residual culture medium at the base was taken out and washed, and the protothallus mass was treated in the following two ways: ① mixed and stirred with 200mL 1 / 2MS salt solution (1 / 2MS culture medium salt without organic matter) for 5s, and then poured into plastic boxes containing sterilized peat soil-perlite, peat soil-vermiculite, river sand-perlite, and peat soil-river sand-perlite matrix, and the combination of all matrices had a dosage ratio of 1:1; ② directly planted in a culture bottle containing sterilized vermiculite.

[0066] Inoculate 2-3 protothallus groups (about 10g) each time, repeat 3 times. For treatment ①, spray the Japanese garden formula solution (calcium nitrate tetrahydrate 945mg / L + potassium nitrate 809mg / L + diammonium phosphate 153mg / L + magnesium sulfate heptahydrate 493mg / L) every other day; for treatment ②, pour 1 / 2MS salt solution into the culture bottle once and tighten the lid without further treatment. Cultivate at 27℃ during the day and 25℃ at night. Transformation is successful when the first true leaf grows. Record the earliest formation of sporophyte, and calculate the transformation rate after 60 days.

[0067] The results are shown in Table 6 below, which show that the time and conversion rate of spores formed by the prothallus of Cibotium chinense in different substrates are significantly different. The mixed substrate spore formation time is the shortest, which is 30 days; the river sand-perlite and vermiculite (in the bottle) substrate spore formation time is the longest, which is 52 days. The peat soil-vermiculite substrate spore conversion rate is the highest, which is 84.42%, and the vermiculite substrate in the bottle has the lowest conversion rate, which is only 26.39%.

[0068] Table 6 Effects of different substrates on the transformation of Cibotium spores (n=3)

[0069]

[0070] Note: The data in the table are mean ± standard deviation. Different lowercase letters in the same column indicate significant differences (P < 0.05).

[0071] The results of using different substrates to induce sporophytes are as follows Figure 3 As shown in FIG. 1 , it is shown that the induced sporophyte transformation effect of peat soil-vermiculite is the best; the transformation effect after 60 days is as shown in FIG. Figure 4 As shown, there was no significant difference in the time for inducing sporophyte formation between peat soil-vermiculite and peat soil-river sand-perlite, while the sporophyte conversion rate of peat soil-vermiculite was significantly higher than that of peat soil-river sand-perlite; therefore, peat soil-vermiculite was selected as the spore transformer matrix for the incomplete tissue culture method to improve the sporophyte conversion rate, shorten the time for seedling formation and acclimatization, accelerate its reproduction and improve the reproduction rate at the same time, and the operation requirements are not strict, which solves the technical problems of seedlings being easily contaminated, fragile and having a low seedling rate in tissue culture bottles.

[0072] Example 4 Sporophyte Transplantation of Cibotium caninum

[0073] The sporophyte to be treated grows 2 to 3 true leaves. When the plant height is 3 to 5 cm, take it out, carefully remove the protothallus blocks remaining at the base, separate it into individual sporophytes, avoid the protothallus affecting the absorption of sporophyte nutrients, and transplant it into a peat soil-vermiculite-perlite matrix. Water it once every 2 days to maintain humidity, cover it with a plastic cover for shade, and remove the cover for routine management after two weeks. Transfer the sporophyte seedlings to the peat soil-vermiculite-perlite matrix and continue to cultivate for 2 weeks. Statistic the survival rate, survival rate % = number of surviving seedlings / total number of transplants × 100%.

[0074] The results are as follows Figure 5 The results showed that the young sporophyte seedlings after transplantation could grow healthily with a survival rate of 90%. Figure 5 As shown in A, the young sporophyte seedlings that have grown for a period of time are Figure 5 As shown in B, a single sporophyte growing in the substrate Figure 5As shown in C, it is shown that all young sporophyte seedlings can grow healthily. The method of the present invention can accelerate the spore propagation of Cibotium chinense and the domestication of sporophytes, and can obtain a large number of sporophyte seedlings in a short period of time. The present invention adopts an incomplete tissue culture method to improve the sporophyte transformation rate, shorten the time for seedling hardening and domestication, accelerate its reproduction and improve the reproduction rate at the same time, and has no strict requirements on operation, thereby solving the technical problems that seedlings are easily contaminated, fragile, and have a low seedling rate in tissue culture bottles.

[0075] The above embodiments are preferred implementation modes of the present invention, but the implementation modes of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications that do not deviate from the spirit and principles of the present invention should be equivalent replacement methods and are included in the protection scope of the present invention.

Claims

1. A prothallus proliferation culture medium for Cibotium caninum, characterized in that: The 1 / 2MS culture medium is used as the basic culture medium and further contains the following components: 10-30 g / L sucrose + 5-7 g / L agar + 50-200 g / L natural organic additive + 0.1 g / L activated carbon; the natural organic additive is banana juice, potato juice or coconut juice.

2. The proliferation medium according to claim 1, characterized in that The components of the 1 / 2MS culture medium are: macroelements KNO3 950mg / L, NH4NO3 825mg / L, MgSO4 90.35mg / L, KH2PO485mg / L; trace elements MnSO4·H2O16.9mg / L, ZnSO4·7H2O 8.6mg / L, H3BO3 6.2mg / L, KI 0.83mg / L, Na2MoO4·2H2O 0.25mg / L, CoCl2·6H2O 0.025mg / L, CuSO4·5H2O0.025mg / L; iron salt FeNaEDTA 36.7mg / L; organic matter calcium salt CaCl2 166.1mg / L; Glycine 2.0mg / L; Thiamine hydrochloride 0.1mg / L; Pyridoxine hydrochloride 0.5mg / L; Nicotinic acid 0.5mg / L; Inositol 100mg / L.

3. The proliferation medium according to claim 1, characterized in that The basic culture medium is 1 / 2MS culture medium, and also contains the following components: 30g / L sucrose + 6g / L agar + 100g / L natural organic additives + 0.1g / L activated carbon.

4. The proliferation medium according to claim 3, characterized in that The natural organic additive is banana juice.

5. Use of the prothallus proliferation medium according to any one of claims 1 to 4 in the rapid propagation of Cibotium caninum.

6. A method for rapid propagation of Cibotium barometz, characterized in that: After the spores of Cibotium chinense are germinated and cultured, they are inoculated into the prothallus proliferation culture medium according to any one of claims 1 to 4 for proliferation culture, and the prothallus after proliferation is transferred to a substrate for indoor culture and then transplanted to obtain the product.

7. The method according to claim 6, characterized in that: The specific method is as follows: S1. Take the spore powder of Cibotium caninum, disinfect it, suspend it with sterile water, and then inoculate it in a solid culture medium for germination culture; S2. The young prothallus clusters with a growth time of more than 30 days in S1 are divided into small prothallus clusters, inoculated into the proliferation medium according to any one of claims 1 to 4, and cultured for proliferation to obtain prothallus clusters after proliferation; S3. After the proliferated prothallus mass is sheared, it is transferred to a sterilized peat soil-vermiculite matrix for further cultivation to obtain a sporophyte seedling; S4. Transplant the sporophyte seedlings into a peat soil-vermiculite-perlite matrix and culture them in a moisturizing manner. After two weeks, culture them in a conventional manner to obtain the young sporophyte seedlings of Cibotium barometz.

8. The method according to claim 7, characterized in that: In step S1, the solid culture medium is selected from 1 / 2MS solid, 1 / 4MS solid or 1 / 4MS solid + 1% sucrose; the culture conditions are: fluorescent light source, temperature 25-27° C., and illumination time 12 h / d.

9. The method according to claim 7, characterized in that: The shearing treatment in step S1 is as follows: mixing the protothallus mass with 1 / 2MS salt solution, stirring and shearing for 5 seconds; the dosage ratio of the peat soil-vermiculite matrix is ​​1:

1.

10. The method according to claim 7, characterized in that: In step S4, the sporophyte seedlings are 3 to 5 cm in height and have 2 to 3 true leaves.

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