Rooting medium and its application in culturing sugarcane plantlets

Abstract

The application belongs to the technical field of plant tissue culture, and discloses a rapid rooting culture medium for sugarcane tissue culture seedlings and a preparation method. The rooting culture medium comprises the following components in a solution with a pH value of 5.8: 1.1g of MS culture medium, 30g of sucrose, 0.5-1.5mg of NAA, 1.0-3.0mg of 6-BA, 1.0-3.0mg of ZT, 1.0-3.0mg of inositol, 0.1-0.2mg of VB1 and 6g of agar. The culture medium of the application is used for culturing sugarcane explants, and has good rooting effect, longer root length and more roots. The culture medium can obviously improve the rooting number, root length and stem length of the sugarcane explants, and significantly improve the rooting rate and transplanting survival rate of the explants.

Description

Technical Field

[0001] This invention pertains to plant tissue culture technology. Background Technology

[0002] Sugarcane (Saccharum officinarum L.) is a tall, solid perennial herb with a robust and well-developed rhizome. The long-standing practice of asexual propagation for seed saving in sugarcane production has led to varietal degeneration, reduced yield, and decreased quality. Through shoot tip culture in tissue culture, viral diseases such as mosaic virus and ratoon dwarfism can be effectively eliminated, preventing varietal degeneration, purifying and rejuvenating the plant, and enhancing sugarcane yield and sucrose content.

[0003] The contamination rate and survival rate of virus-free sugarcane seedlings, as well as the survival rate of transplanted seedlings, are important factors affecting the cost of sugarcane production. Currently, sugarcane tissue culture propagation is fast, but explant rooting efficiency is low, rooting time is long, and general culture media have low efficiency. Therefore, improving and innovating sugarcane rooting culture media to increase explant rooting rate and survival rate is an urgent need in the sugarcane production process. Summary of the Invention

[0004] The purpose of this invention is to provide a rooting medium comprising the following components per liter of solution at pH 5.8: 1.1 g MS medium, 30 g sucrose, 0.5 to 1.5 mg NAA, 1.0 to 3.0 mg 6-BA, 1.0 to 3.0 mg ZT, 1.0 to 3.0 mg inositol, 0.1 to 0.2 mg VB1, and 6 g agar.

[0005] Furthermore, the NAA is 0.5 mg, 1.0 mg, or 1.5 mg.

[0006] Furthermore, the 6-BA is 1.0 mg, 2.0 mg, or 3.0 mg.

[0007] Furthermore, the ZT is 1.0 mg, 2.0 mg, or 3.0 mg.

[0008] Furthermore, the inositol is 1.0 mg, 2.0 mg, or 3.0 mg.

[0009] Furthermore, the VB1 is 0.1 mg or 0.2 mg.

[0010] The present invention also provides the application of the rooting medium in the cultivation of sugarcane tissue culture seedlings.

[0011] The culture medium of this invention produces excellent rooting results in sugarcane explants, with longer and more numerous roots. It significantly improves the issues of root number and root length in sugarcane explants, and substantially increases the rooting rate and transplant survival rate. Attached Figure Description

[0012] Figure 1 These are images of rootstocks cultured in the control group. A is a bottom view, and B is a front view.

[0013] Figure 2 This is a diagram of a group of cultured root seedlings. A is a bottom view, and B is a front view.

[0014] Figure 3 These are two sets of images showing the cultured rootstocks. A is a bottom view, and B is a front view.

[0015] Figure 4 These are images of three groups of cultured seedlings. A is a bottom view, and B is a front view. Detailed implementation method:

[0016] Unless otherwise specified, the sugarcane variety used in the following examples is Guitang 42. The technical solution of the present invention will be further illustrated below through specific embodiments.

[0017] Using healthy Guitang 42 sugarcane tissue culture seedlings from the same batch, ineffective tillers and yellowed leaves were removed. The clustered buds at the differentiation stage of the sugarcane tissue culture seedlings were used as raw materials, and the clustered buds were divided into individual plants, which were then inoculated onto a modified rooting medium. The control medium was a commonly used rooting medium.

[0018] Example 1

[0019] 1. Preparation of culture medium

[0020] (1) Conventional rooting medium (control):

[0021] MS medium: 4.4 g / L, 30 g / L sucrose, 6 g / L agar, 1.5 mg / L NAA (naphthaleneacetic acid), pH 5.8.

[0022] The preparation method is as follows: Add sucrose to distilled water, then add MS medium and NAA according to the above ratio, mix in agar, bring the volume to 1L with distilled water, adjust the pH to 5.8 with NaOH or HCl, dispense into culture flasks, autoclave at 121℃ for 15min, and cool to obtain the rooting medium.

[0023] (2) Improved rooting medium (treatment 1):

[0024] MS medium: 1.1 g / L, 30 g / L sucrose, 0.50 mg / L NAA (naphthaleneacetic acid), 1.0 mg / L 6-BA (6-benzylaminopurine), 1.0 mg / L ZT (zeatin), 1.0 mg / L inositol, 0.10 mg / L VB1 (vitamin B1), 6 g / L agar, pH 5.8.

[0025] The preparation method is as follows: Add sucrose to distilled water, and then add MS medium, NAA, 6-BA, ZT, inositol and VB1 according to the above ratio. Mix them into agar, bring the volume to 1L with distilled water, adjust the pH to 5.8 with NaOH or HCl, dispense into culture flasks, autoclave at 121℃ for 15min, and cool to obtain the modified rooting medium.

[0026] 2. Cultivation of clustered buds:

[0027] Sugarcane explant shoot clusters, after proliferation culture, were divided into single shoot clusters and inoculated into rooting medium. The culture was then maintained at 24°C with 16 hours of light and 70 μmol / L phosphate. -2 s -1 Light intensity was applied for 30 days. The rooting effects of the two culture media were compared. Ten groups (bottles) were set up as the control group, with 10 plants in each group (bottle). The number of roots and root length (in cm) of each plant were measured.

[0028] Figure 1 These are images of rootstocks cultured in the control group. In the image, A is a bottom view and B is a front view. Figure 2 This is a diagram of a group of cultured root seedlings. A is a bottom view, and B is a front view.

[0029] Table 1. Values ​​of root number and average root length

[0030]

[0031]

[0032]

[0033] Example 2

[0034] 1. Preparation of culture medium

[0035] (1) Conventional rooting medium (control): MS medium 4.4 g / L, 30 g / L sucrose, 6 g / L agar, 1.5 mg / L NAA, pH 5.8.

[0036] The preparation method is as follows: Add sucrose to distilled water, then add MS medium and NAA according to the above ratio, mix in agar, bring the volume to 1L with distilled water, adjust the pH to 5.8 with NaOH or HCl, dispense into culture flasks, autoclave at 121℃ for 15min, and cool to obtain the conventional rooting medium.

[0037] (2) Rooting medium (treatment 2): MS medium 1.1 g / L, 30 g / L sucrose, 1.0 mg / L NAA, 2.0 mg / L 6-BA, 2.0 mg / L ZT, 2.0 mg / L inositol, 0.10 mg / L VB1, 6 g / L agar, pH 5.8.

[0038] The preparation method is as follows: Add sucrose to distilled water, and then add MS medium, NAA, 6-BA, ZT, inositol and VB1 according to the above ratio. Mix them into agar, bring the volume to 1L with distilled water, adjust the pH to 5.8 with NaOH or HCl, dispense into culture flasks, autoclave at 121℃ for 15min, and cool to obtain the rooting medium.

[0039] 2. Cluster bud culture. The cluster buds of sugarcane explants, after proliferation culture, were divided into single-plant cluster buds and inoculated onto rooting medium. The culture was then maintained at 24℃ with 16 hours of light and 70 μmol / L. -2 s -1 Light intensity was applied for 30 days. The rooting effects of the two culture media were compared. Ten groups (bottles) were set up as the control group, with 10 plants in each group (bottle). The number of roots and root length (in cm) of each plant were measured.

[0040] Figure 3 These are two sets of images showing the cultured rootstocks. A is a bottom view, and B is a front view.

[0041] Table 2. Root Count and Average Root Length

[0042]

[0043]

[0044]

[0045]

[0046] Example 3

[0047] 1. Preparation of culture medium

[0048] (1) Conventional rooting medium (control): MS medium 4.4 g / L, 30 g / L sucrose, 6 g / L agar, 1.5 mg / L NAA, pH 5.8.

[0049] The preparation method is as follows: Add sucrose to distilled water, then add MS medium and NAA according to the above ratio, mix in agar, bring the volume to 1L with distilled water, adjust the pH to 5.8 with NaOH or HCl, dispense into culture flasks, autoclave at 121℃ for 15min, and cool to obtain the conventional rooting medium.

[0050] (2) Rooting medium (treatment 3): MS medium 1.1 g / L, 30 g / L sucrose, 1.50 mg / L NAA, 3.0 mg / L 6-BA, 3.0 mg / L ZT, 3.0 mg / L inositol, 0.20 mg / L VB1, 6 g / L agar, pH 5.8.

[0051] The preparation method is as follows: Add sucrose to distilled water, and then add MS medium, NAA, 6-BA, ZT, inositol and VB1 according to the above ratio. Mix them into agar, bring the volume to 1L with distilled water, adjust the pH to 5.8 with NaOH or HCl, dispense into culture flasks, autoclave at 121℃ for 15min, and cool to obtain the rooting medium.

[0052] 2. Cluster bud culture. Cluster buds from sugarcane explants that have undergone propagation culture are divided into single-plant cluster buds, inoculated onto rooting medium, and cultured at 24℃ with 16 hours of light and 70 μmol / L. -2 s -1 Light intensity was measured, and the culture was carried out for 30 days. The rooting effects of the two culture media were compared. Ten groups (bottles) were set up in the control group, with 10 plants in each group (bottle). The number of roots and root length (in cm) of each plant were measured.

[0053] Figure 4 These are images of three groups of cultured seedlings. A is a bottom view, and B is a front view.

[0054] Table 3. Root Count and Average Root Length

[0055]

[0056]

[0057]

[0058] The survival rate of transplanted seedlings in the control group and treatment groups 1 to 3 was statistically analyzed. For treatments with the same total number of seedlings after transplanting, the number of roots, root length, and other data were also statistically analyzed. The results are as follows:

[0059] Table 4 Comparison of Rooting Culture Medium Results

[0060] Comparison 10 10 14.36±2.21 0.87±0.46 88 Process 1 10 10 19.99±2.58 1.58±0.21 94 Process 2 10 10 27.7±2.60 1.91±0.38 98 Process 3 10 10 21.97±2.68 1.39±0.21 96

[0061] The single-plant clusters cultivated using the culture medium of this invention exhibit excellent rooting performance, with longer and more numerous roots. This significantly improves upon the shortcomings of existing sugarcane explants, which have fewer roots and shorter, thinner roots, thereby substantially increasing the rooting rate and transplant survival rate of the explants.

Claims

1. A rooting culture medium, characterized in that, The solution at pH 5.8 contains the following components per liter: 1.1 g MS medium, 30 g sucrose, 0.5 to 1.5 mg NAA, 1.0 to 3.0 mg 6-BA, 1.0 to 3.0 mg ZT, 1.0 to 3.0 mg inositol, 0.1 to 0.2 mg VB1, and 6 g agar.

2. The rooting medium according to claim 1, characterized in that the NAA is 0.5 mg, 1.0 mg or 1.5 mg.

3. The rooting culture medium according to claim 1, characterized in that the 6-BA is 1.0 mg, 2.0 mg or 3.0 mg.

4. The rooting culture medium according to claim 1, characterized in that the ZT is 1.0 mg, 2.0 mg or 3.0 mg.

5. The rooting culture medium according to claim 1, characterized in that the inositol is 1.0 mg, 2.0 mg or 3.0 mg.

6. The rooting culture medium according to claim 1, characterized in that the VB1 is 0.1 mg or 0.2 mg.

7. The application of the rooting medium as described in any one of claims 1 to 6 in the cultivation of sugarcane tissue culture seedlings.

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