A method for breeding and culture medium of tagetes erecta

Abstract

The present application relates to the technical field of plant culture, and specifically discloses a method for breeding tagetes and a culture medium.The method for breeding tagetes comprises the step of culturing explants of tagetes into rooted seedlings;an induction culture medium used in the culture of the explants comprises quercetagetin, 6-benzylaminopurine and naphthaleneacetic acid;wherein the mass ratio of quercetagetin, 6-benzylaminopurine and naphthaleneacetic acid is (10-20):(0.1-1):(0.01-0.1);and / or a rooting culture medium used in the culture of the explants comprises quercetagetin and naphthaleneacetic acid;wherein the mass ratio of quercetagetin and naphthaleneacetic acid is (10-20):(0.01-0.1).The method can obtain a large number of tagetes seedlings in a short time, is simple to operate, has a short culture period, is low in input cost, high in propagation efficiency, and suitable for large-scale production and application.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of plant culture, in particular to a method for breeding Tagetes and a culture medium. BACKGROUND

[0002] Tagetes is an annual herb of the genus Tagetes in the family Asteraceae, mainly divided into two categories of pigment Tagetes and ornamental Tagetes, the former has large flower quantity, long flowering period and rich lutein in flowers. Currently, commercial Tagetes seeds are mainly obtained by hybridization, and due to the selfing of parents for many years, the degeneration is serious, the yield is reduced, and the popular planting of Tagetes is seriously hindered. At present, the asexual rapid propagation technology is widely used in plant breeding and cultivation fields, which not only can maintain the original excellent traits of parents, but also can obtain a large number of genotype stable seedlings in a short time, and can provide valuable materials for Tagetes breeding.

[0003] The traditional asexual propagation methods mainly include cutting, grafting, division and layering, and the propagation efficiency and survival rate are low. In recent years, plant tissue culture technology has developed rapidly, but there are problems such as complex culture medium composition, long tissue culture period, low induction rate and high cost. At present, the research on asexual rapid propagation technology of Tagetes is still relatively few, mainly focusing on the establishment of tissue culture system, which has great technical difficulty and is prone to vitrification problem, and is difficult to scale application and promotion. Therefore, it is necessary to further study the plant tissue culture technology of Tagetes. SUMMARY

[0004] One of the purposes of the present application is to provide a method for breeding Tagetes with high propagation efficiency.

[0005] The present application provides a method for breeding Tagetes, which comprises the step of culturing explants of Tagetes into rooted seedlings; the induction culture medium used in the culture of the explants comprises quercetagetin, 6-benzylaminopurine and naphthaleneacetic acid; wherein the mass ratio of quercetagetin, 6-benzylaminopurine and naphthaleneacetic acid is (10-20):(0.1-1):(0.01-0.1).

[0006] And / or, the rooting culture medium used in the culture of the explants comprises quercetagetin and naphthaleneacetic acid; wherein the mass ratio of quercetagetin and naphthaleneacetic acid is (10-20):(0.01-0.1).

[0007] The present application researches and finds that the use of quercetagetin can improve the breeding effect when breeding Tagetes, specifically, it is found that when buds are induced and roots are induced, the combination of quercetagetin with other specific components in the culture medium can promote the growth of buds and roots, significantly improve the induction rate of buds and roots, lay a good foundation for the field planting of plants, and obtain field plants with better growth conditions.

[0008] Preferably, the induction medium and rooting medium of the present application are used in the culture of explants.

[0009] The present application can obtain a large number of gazania seedlings in a short time, and the culture period is greatly shortened compared with the method which needs to go through the culture process of dedifferentiation and redifferentiation.

[0010] The present application can obtain a large number of gazania hybrid lines or other process materials with consistent genetic background, which not only saves the time cost of artificial crossing pollination in breeding, but also shortens the breeding period.

[0011] In the gazania breeding method of the present application, the induction medium further comprises MS inorganic salt, sucrose and plant gel; and the rooting medium further comprises MS inorganic salt, sucrose and plant gel.

[0012] In the present application, the MS inorganic salt is a powder base used in the preparation of conventional MS medium in the art.

[0013] The plant gel is a commercially available product commonly used in plant tissue culture in the art.

[0014] In the gazania breeding method of the present application, preferably, the induction medium comprises 4.94 g / L MS inorganic salt, 0.01-0.02 g / L quercetin, 0.1-1 mg / L 6-benzylaminopurine, 0.01-0.1 mg / L naphthalene acetic acid, 20-30 g / L sucrose and 5-10 g / L plant gel, and the pH value is 5.8-6.

[0015] The rooting medium comprises 2.47 g / L MS inorganic salt, 0.01-0.02 g / L quercetin, 0.01-0.1 mg / L naphthalene acetic acid, 20-40 g / L sucrose, 5-10 g / L plant gel, and the pH value is 5.8-6.

[0016] In the gazania breeding method of the present application, the culture of the explants comprises inoculating the stem segments with bud points into the induction medium, culturing for 15-20 days to obtain young buds, and then transferring the young buds to the rooting medium for 15-30 days to obtain rooted seedlings.

[0017] In the gazania breeding method of the present application, the height of the young buds is 3-5 cm.

[0018] And / or, the culture conditions are temperature 25±2℃, humidity 60-85%, light intensity 6000-8000 LUX, light time per day 14-16 h, and dark time 10-8 h.

[0019] The marigold breeding method of the present application further comprises the steps of acclimatizing and transplanting the rooted seedlings; the rooted seedlings used in acclimatization have a height of 8-10 cm and 3-5 roots, and the acclimatization conditions are as follows: temperature 25±2℃; humidity 60-85%; light intensity 6000-8000 LUX; light time per day 14-16h, dark time 10-8h, acclimatization time 7-10d, and the substrate comprises 1 part of perlite, 1 part of vermiculite and 3-5 parts of grass carbon.

[0020] The acclimatization and transplanting method of the present application is beneficial to improving the survival rate of marigold seedlings after field transplanting.

[0021] In the marigold breeding method of the present application, the explants are obtained from the rooted seedlings or field plants obtained by tissue culture; if the explants are obtained from the field plants, a sterilization step is further included before culture.

[0022] As a specific embodiment, the efficient rapid propagation method of marigold of the present application comprises the following steps:

[0023] S1, preparation of culture medium: preparation of induction culture medium and rooting culture medium;

[0024] S2, inoculation of explants: select tender stem segments with bud points as explants, which can be obtained from the rooted seedlings in S3 or field plants, and the stem segments obtained from the field plants need to be subjected to surface sterilization treatment;

[0025] S3, bud induction and rooting: inoculate the tender stem segments with bud points into the induction culture medium, culture for 15-20d, then select appropriate size buds and transfer them to the rooting culture medium for culture for 15-30d to obtain the rooted seedlings, which can be used to continue to provide the stem segments with bud points in S2 or directly enter S4;

[0026] S4, acclimatization and transplanting: transplant the rooted seedlings into the soil of the seedling substrate, and acclimatize for 7-10d before field transplanting.

[0027] The present application starts from the culture of stem segments with bud points, and the stem segments with bud points obtained in S3 can be used to continue step S2, so that the propagation efficiency can be greatly improved without affecting the growth of seedlings, and a large number of marigold seedlings can be obtained in a short time.

[0028] The present application further provides an induction culture medium, which is as described above.

[0029] The induction culture medium of the present application can improve the bud induction rate of marigold explants during culture, and lay an ideal foundation for subsequent tissue culture. It can be used with other known rooting culture media in the art.

[0030] The application also provides a rooting medium, which is as described above.

[0031] The induction medium of the application can improve the root induction rate of the gerbera explant culture, and provide more ideal materials for subsequent field planting. The induction medium can be used in combination with other known bud induction media in the art.

[0032] The application also provides application of the above induction medium and / or rooting medium in gerbera breeding.

[0033] The application has at least the following beneficial effects:

[0034] The gerbera breeding method provided by the application can obtain a large number of gerbera seedlings in a short time. The whole process is simple in operation, short in culture period, low in input cost, and high in breeding efficiency, and is suitable for large-scale production and application. DETAILED DESCRIPTION

[0035] The preferred embodiments of the application will be described in detail below with reference to the examples. It should be understood that the following examples are given only for the purpose of illustration and are not intended to limit the scope of the application. Those skilled in the art can make various modifications and replacements to the application without departing from the spirit and principles of the application.

[0036] In the following examples, the experimental methods are conventional unless otherwise specified. In the following examples, the materials, reagents, etc. are commercially available or prepared according to conventional methods in the art unless otherwise specified. The main component of the plant gel used in the specific embodiment part of the application is gellan gum, which is purchased from Kangbeisi Biological, CAS No.: 71010-52-1.

[0037] Example 1

[0038] The present embodiment provides a method for gerbera tissue culture, which comprises the following steps:

[0039] S1, preparation of the culture medium:

[0040] Twelve kinds of induction media (pH 5.8) were prepared with water, including 4.94 g / L MS inorganic salt (solid powder), 25 g / L sucrose, 8 g / L plant gel (purchased from Kangbeisi Biological, CAS No.: 71010-52-1), and quercetagetin, 6-benzylaminoadenine and / or naphthalene acetic acid. The specific concentrations of quercetagetin, 6-benzylaminoadenine and naphthalene acetic acid in each induction medium are shown in Table 1. The induction media numbered 1-9 are comparative schemes, and the induction media numbered 10-12 are the schemes of the present application.

[0041] The rooting medium was configured with water, including: 2.47 g / L MS inorganic salt (solid powder), 25 g / L sucrose, 8 g / L plant gel, pH 5.8.

[0042] S2, inoculation of explants: select the bud point of the tender stem segment of the color tagetes "Crimson No. 1" field plant as the explant (the total number is 50), and the stem segment needs to be surface sterilized before inoculation.

[0043] The surface sterilization method is 75% alcohol immersion treatment for 30S, sterile water washing 3-4 times, 1 min each time, 5% sodium hypochlorite solution treatment for 10 min, sterile water washing 4-5 times, 1 min each time.

[0044] S3, bud induction and rooting: the bud point of the tender stem segment is inoculated into different induction medium, and cultured at 25±2℃, humidity 60-85%, light intensity 7000LUX, light time 15h per day for 15d, then the height of the young bud is selected 3-5cm, and it is transferred to the rooting medium under the same conditions as above for 15d, and the rooting seedlings are obtained. The rooting seedlings can provide the stem segment with bud points for S2 or directly enter S4.

[0045] The effect of different induction medium on bud induction rate is counted, and the bud induction rate = the number of induced buds / the total number of inoculated explants x 100%, and the results are shown in Table 1.

[0046] Table 1

[0047]

[0048] S4, acclimation and transplanting: the rooting seedlings with a height of 8-10cm and 3-5 roots are transplanted into the seedling substrate soil (1 part of perlite, 1 part of vermiculite, 4 parts of grass carbon soil), and the culture conditions of step S3 are acclimated for 10d, and then the field transplanting is carried out.

[0049] The field transplanting results of each combination are counted, and the transplanting survival rate is more than 90%, and there is no difference between each combination, and the statistical number of each combination is 100 plants. The growth of the plants after transplanting is good.

[0050] Comparative example 1

[0051] This comparative example provides a method for tissue culture of tagetes, which is basically the same as the method of example 1, and the only difference is that the naphthalene acetic acid in the No. 11 induction medium is replaced by indole-3-acetic acid. The bud induction rate is 60%. The transplanting survival rate is 92%.

[0052] Comparative example 2

[0053] The comparative example provides a method for carrying out tissue culture of Tagetes, which is basically the same as the method of Example 1, with the only difference being that 6-benzylaminopurine is replaced by kinetin in the No. 11 induction medium. The bud induction rate is 70%. The transplanting survival rate is 90%.

[0054] Comparative Example 3

[0055] The comparative example provides a method for carrying out tissue culture of Tagetes, which is basically the same as the method of Example 1, with the only difference being that the concentration of quercetagetin is 0.03 g / L, the concentration of 6-benzylaminopurine is 0.1 mg / L, and the concentration of naphthalene acetic acid is 0.2 mg / L. The bud induction rate is 50%, and the transplanting survival rate is 91%.

[0056] Example 2

[0057] The example provides a method for carrying out tissue culture of Tagetes, which comprises the following steps:

[0058] S1, preparation of the medium:

[0059] The induction medium is prepared with water, which comprises: 4.94 g / L MS inorganic salt (solid powder), 0.01 g / L quercetagetin, 0.1 mg / L 6-benzylaminopurine, 0.01 mg / L naphthalene acetic acid, 20 g / L sucrose, 5 g / L plant gel, and pH 5.8.

[0060] The rooting medium (pH 5.8) is prepared with water, which comprises: 2.47 g / L MS inorganic salt (solid powder), 25 g / L sucrose, 8 g / L plant gel, and quercetagetin, 6-benzylaminopurine and / or naphthalene acetic acid. The specific concentrations of quercetagetin, 6-benzylaminopurine and naphthalene acetic acid in each rooting medium are shown in Table 2. The rooting media numbered 4-12 are comparative schemes, and the rooting media numbered 1-3 are the schemes of the present application.

[0061] S2, inoculation of the explants: the tender stem segments with buds of the pigment Tagetes "Crimson No. 1" field plants are selected as the explants (the total number is 50), and the stem segments need to be subjected to surface sterilization treatment before inoculation.

[0062] The method for surface sterilization is 75% alcohol immersion treatment for 30 s, sterile water washing for 3-4 times, each time for 1 min, 5% sodium hypochlorite solution treatment for 10 min, and sterile water washing for 4-5 times, each time for 1 min.

[0063] S3, bud induction and rooting: the young stem segments with bud points are inoculated on the induction medium, and cultured at a temperature of 25±2℃, a humidity of 60-85%, a light intensity of 7000 LUX, and a light time of 15h per day for 15d, after which the young buds with a height of 3-5cm are transferred to different rooting media and cultured under the same conditions for another 15d to obtain the rooted seedlings, which can be used to provide the stem segments with bud points for S2 or directly enter S4.

[0064] The effects of different types of rooting media on the root induction rate are counted, the root induction rate being the number of roots with a length of more than 3cm / total number of roots x 100% after 7d of culture, and the results are shown in Table 2.

[0065] Table 2

[0066]

[0067] S4, acclimatization and transplanting: the rooted seedlings with a height of 8-10cm and a number of roots of 3-5 are transplanted into the seedling substrate soil (1 part of perlite, 1 part of vermiculite, and 4 parts of grass carbon soil), and acclimatized for 10d under the culture conditions of S3 to be transplanted into the field.

[0068] The field transplanting results of each combination are counted, and the transplanting survival rates are all above 90%, with no difference among the combinations, and the number of plants counted for each combination is 100. The plants after transplanting grow well.

[0069] Comparative Example 4

[0070] This comparative example provides a method for tissue culture of gazania, which is basically the same as that of Example 2, except that the naphthaleneacetic acid in the No. 2 rooting medium is replaced by indole-3-acetic acid. The root induction rate is 75%, and the transplanting survival rate is 93%.

[0071] Comparative Example 5

[0072] This comparative example provides a method for tissue culture of gazania, which is basically the same as that of Example 2, except that the concentration of quercetagetin is 0.025g / L and the concentration of naphthaleneacetic acid is 0.1mg / L. The root induction rate is 76%, and the transplanting survival rate is 92%.

[0073] Example 3

[0074] This example provides a method for tissue culture of gazania, which comprises the following steps:

[0075] S1, Preparation of medium: First, the induction medium is prepared with water, which includes: 4.94 g / L MS inorganic salt (solid powder), 0.01 g / L quercetin, 0.1 mg / L 6-benzylaminopurine, 0.01 mg / L naphthalene acetic acid, 20 g / L sucrose, 5 g / L plant gel, pH 5.8;

[0076] Then, the rooting medium is prepared with water, which includes: 2.47 g / L MS inorganic salt (solid powder), 0.01 g / L quercetin, 0.01 mg / L naphthalene acetic acid, 20 g / L sucrose, 5 g / L plant gel, pH 5.8.

[0077] S2, Inoculation of explants: the tender stem segments with bud points of the pigment tagetes "Crimson No. 1" are selected as explants, which can be obtained from the rooted seedlings in the step S3 and the field plants, and the stem segments obtained from the field plants need to be subjected to surface disinfection treatment (for details, see Example 1).

[0078] S3, bud induction and rooting: the tender stem segments with bud points are inoculated onto the induction medium, and are cultured at a temperature of 25±2℃, a humidity of 60-85%, a light intensity of 7000 LUX, and a light time of 15 h per day for 15 d, after which the young buds with a height of 3-5 cm are selected and transferred to the rooting medium for further culture under the same conditions for 15 d to obtain the rooted seedlings, which can be used to provide the stem segments with bud points for S2 or directly used in S4.

[0079] S4, acclimatization and transplanting: the rooted seedlings with a height of 8-10 cm and 3-5 roots are transplanted into the seedling substrate soil (for details, see Example 1), and are acclimatized for 10 d under the culture conditions of step S3, and then are transplanted into the field.

[0080] The growth data of the tagetes seedlings obtained by the above method and the seedling seedlings obtained by conventional seed propagation are counted, and the results are shown in Table 3.

[0081] Table 3

[0082]

[0083] a: the tagetes seedlings obtained by the method of the present application, b: the seedling seedlings obtained by conventional seed propagation, the counting period is the present budding period, and each group has 10 seedlings.

[0084] Example 4

[0085] The embodiment provides a method for carrying out tissue culture of Tagetes, which is basically the same as the method in Embodiment 3, and the difference is that the induction medium comprises 4.94 g / L MS inorganic salt (solid powder), 0.02 g / L quercetin, 1 mg / L 6-benzylaminopurine, 0.1 mg / L naphthalene acetic acid, 30 g / L sucrose, 10 g / L plant gel, and the pH value is 5.8.

[0086] The rooting medium comprises 2.47 g / L MS inorganic salt (solid powder), 0.02 g / L quercetin, 0.1 mg / L naphthalene acetic acid, 40 g / L sucrose, 10 g / L plant gel, and the pH value is 5.8.

[0087] The growth data of the Tagetes seedlings obtained by using the above method and the seedlings obtained by using conventional seed propagation are counted, and the results are shown in Table 4.

[0088] Table 4

[0089]

[0090] a is the Tagetes seedling obtained by using the method, b is the seedling obtained by using conventional seed propagation, and the counting period is the present budding period, and each group has 10 seedlings.

[0091] Embodiment 5

[0092] The efficient rapid propagation method of the Tagetes in the embodiment is applied to four different strains of pigment Tagetes "Xianghong No. 1", "Guanying", ornamental Tagetes "Jili", and "Taishan".

[0093] The specific method is as follows:

[0094] S1, preparation of the medium: first, the induction medium is prepared with water, and the induction medium comprises 4.94 g / L MS inorganic salt (solid powder), 0.01 g / L quercetin, 0.1 mg / L 6-benzylaminopurine, 0.01 mg / L naphthalene acetic acid, 20 g / L sucrose, 5 g / L plant gel, and the pH value is 5.8;

[0095] Then, the rooting medium is prepared with water, and the rooting medium comprises 2.47 g / L MS inorganic salt (solid powder), 0.01 g / L quercetin, 0.01 mg / L naphthalene acetic acid, 20 g / L sucrose, 5 g / L plant gel, and the pH value is 5.8.

[0096] S2, inoculation of the explant: the tender stem segments with bud points of the four different types of Tagetes are selected as the explants (the number of the explants of each type of Tagetes is 50), and the stem segments are subjected to surface disinfection treatment (for details, refer to Embodiment 1).

[0097] S3, bud induction and rooting: the young stem segments with bud points were inoculated on the induction medium, and cultured at temperature 25±2℃; humidity 60-85%; light intensity 7000LUX; light time 15h per day for 15d, then the young buds with height 3-5cm were selected and transferred to the rooting medium and cultured for another 15d under the same conditions as above to obtain the rooted seedlings, which can provide the stem segments with bud points for S2 or directly enter S4.

[0098] S4, acclimatization and transplanting: the rooted seedlings with height 8-10cm and root number 3-5 were transplanted into the seedling substrate soil (see Example 1 for details), and acclimatized for 10d under the culture conditions of S3 to be transplanted into the field.

[0099] The data of the whole process were recorded and analyzed, and the bud induction rate, root induction rate and propagation efficiency of the four types of gazania were compared. The propagation efficiency was the number of the finally obtained rooted seedlings / the total number of the inoculated explants, and the results were shown in Table 5. A, B, C and D represented pigment gazania "No.1 scarlet", "Guanying" and ornamental gazania "miracle", "Tayshan", respectively.

[0100] Table 5

[0101]

[0102] The growth traits of the field plants of different strains obtained by this method were counted, and the counting period was the present budding period, and the number of counted seedlings was 10, and the results were shown in Table 6.

[0103] Table 6

[0104]

[0105] Although the present application has been described in detail with general description and specific embodiments above, some modifications or improvements can be made on the basis of the present application, which is obvious to those skilled in the art. Therefore, these modifications or improvements made on the basis of not deviating from the spirit of the present application, all belong to the scope of the present application claimed.

Claims

1. A method for propagating marigolds, characterized in that, The step includes culturing marigold explants into rooted seedlings, the step comprising: Stem segments with buds were inoculated onto an induction medium and cultured to obtain young shoots. The young shoots were then transferred to a rooting medium and cultured to obtain rooted seedlings. The active ingredients in the induction culture medium consist of quercetin, 6-benzylaminopurine, and naphthaleneacetic acid, wherein the mass ratio of quercetin, 6-benzylaminopurine, and naphthaleneacetic acid is (10-20):(0.1-1):(0.01-0.1). The active ingredients in the rooting culture medium consist of quercetin and naphthaleneacetic acid, wherein the mass ratio of quercetin to naphthaleneacetic acid is (10-20):(0.01-0.1).

2. The method for propagating marigolds according to claim 1, characterized in that, The induction medium is composed of active ingredients, MS inorganic salts, sucrose and plant gel, wherein the active ingredients in the induction medium are as described in claim 1; The rooting medium is composed of active ingredients, MS inorganic salts, sucrose and plant gel, wherein the active ingredients in the rooting medium are as described in claim 1.

3. The method for propagating marigolds according to claim 2, characterized in that, The induction medium consisted of: 4.94 g / L MS inorganic salts, 0.01-0.02 g / L quercetin, 0.1-1 mg / L 6-benzylaminopurine, 0.01-0.1 mg / L naphthaleneacetic acid, 20-30 g / L sucrose, and 5-10 g / L plant gel, with a pH of 5.8-6. The rooting medium consisted of 2.47 g / L MS inorganic salts, 0.01-0.02 g / L quercetin, 0.01-0.1 mg / L naphthaleneacetic acid, 20-40 g / L sucrose, and 5-10 g / L plant gel, with a pH of 5.8-6.

4. The method for propagating marigolds according to any one of claims 1-3, characterized in that, The culture time on the induction medium is 15-20 days; the culture time on the rooting medium is 15-30 days.

5. The method for propagating marigolds according to claim 4, characterized in that, The height of the young shoot is 3-5cm; And / or, the culture conditions are: temperature 25±2℃; humidity 60-85%; light intensity 6000-8000LUX; light duration 14-16h and dark duration 10-8h per day.

6. The method for propagating marigolds according to any one of claims 1-3 and 5, characterized in that, It also includes the steps of acclimatizing and transplanting the rooted seedlings; the rooted seedlings used for acclimatization are 8-10cm in height and have 3-5 roots. The acclimatization conditions are: temperature 25±2℃; humidity 60-85%; light intensity 6000-8000LUX; daily light time 14-16h, dark time 10-8h, acclimatization time 7-10d. The substrate by weight includes 1 part perlite, 1 part vermiculite, and 3-5 parts peat moss.

7. The method for propagating marigolds according to claim 4, characterized in that, It also includes the steps of acclimatizing and transplanting the rooted seedlings; the rooted seedlings used for acclimatization are 8-10cm in height and have 3-5 roots. The acclimatization conditions are: temperature 25±2℃; humidity 60-85%; light intensity 6000-8000LUX; daily light time 14-16h, dark time 10-8h, acclimatization time 7-10d. The substrate by weight includes 1 part perlite, 1 part vermiculite, and 3-5 parts peat moss.

8. The method for propagating marigolds according to any one of claims 1-3, 5, and 7, characterized in that, The explants are derived from rooted seedlings obtained through tissue culture or from field plants; if the explants are derived from field plants, a sterilization step is also included before culture.

9. The method for propagating marigolds according to claim 4, characterized in that, The explants are derived from rooted seedlings obtained through tissue culture or from field plants; if the explants are derived from field plants, a sterilization step is also included before culture.

10. The method for propagating marigolds according to claim 6, characterized in that, The explants are derived from rooted seedlings obtained through tissue culture or from field plants; if the explants are derived from field plants, a sterilization step is also included before culture.

11. An induction culture medium, characterized in that, As described in any one of claims 1-3.

12. A rooting culture medium, characterized in that, As described in any one of claims 1-3.

13. The application of the induction medium of claim 11 and / or the rooting medium of claim 12 in the propagation of marigolds.

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