Method for generating Japan lawn grass plant by mature embryo calls induction and culture medium
A basic medium and the technology of zoysia grass, which are applied in the field of plant tissue culture and plant regeneration, can solve the problems of difficult long-term subculture and differentiation ability of callus, and achieve long-term subculture preservation and maintain differentiation ability, The effect of good differentiation ability
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Embodiment 1
[0035] Embodiment 1: the establishment of Japanese zoysia regeneration system
[0036] 1. The source and treatment of test materials:
[0037]Mature Japanese Zoysia (Zoysia japonica Steud) seeds of the test material of the present invention were purchased from Beijing China Seed Industry Group. In the present invention, induction, subculture, and differentiation of callus of Zoysia japonica are not limited by species.
[0038] Grind Zoysia japonica seeds with distilled water in a mortar for about 15 minutes, rinse under tap water to remove the seed coat, and then disinfect with 76% (V / V) alcohol for 30 seconds on an ultra-clean workbench (or sterile inoculation box) , and then sterilized with 0.1% (W / V) mercuric chloride solution for 20-25 minutes, rinsed with sterile water 5 times, and washed for 3-5 minutes each time to obtain sterilized seeds for processing and set aside.
[0039] 2. Culture medium design:
[0040] Table 1 lists the composition and consumption amount of ...
Embodiment 2
[0049] Example 2: Effects of different concentrations of KT on long-term subculture preservation of callus
[0050] (1) Material:
[0051] A callus in a differentiated state formed after 2-3 generations of callus proliferation and culture, and the callus is subjected to long-term subculture.
[0052] (2) Preparation of callus subculture medium:
[0053] The preparation of callus subculture medium is shown in Table 1. In order to explore the effects of different concentrations of KT on the continuously subcultured Zoysia japonica, in this example the applicant specially added the culture medium shown in Table 2 for comparative experiments.
[0054] 3. Training method:
[0055] The induced compact callus formed a callus in a differentiated state after 2-3 generations of subculture (see Figure 4 shown). Select the yellow-white, granular, and well-dispersed callus around the callus in the differentiated state, or the granular, dry, and hard callus formed around the bud struc...
Embodiment 3
[0058] Embodiment 3: The differentiation rate of the Zoysia japonica callus of continuous subculture on different differentiation medium
[0059] The preparation of medium for callus differentiation is shown in Table 1.
[0060] The long-term preserved callus of the present invention often coexists in multiple states (see Figure 5 ), the black arrow points to the callus selected during preservation, the blue arrow points to the callus with only proliferative ability but no differentiation ability, and the red arrow points to a callus with a higher degree of organization , its production is a sign that the subcultured callus of the present invention has preservation value.
[0061] In this example, applicants used the callus that had been subcultured for 6 generations for the experiment of differentiating shoots and plants. On the basis of the general differentiation medium shown in Table 1, the applicant specially designed the differentiation medium shown in Table 3 to dete...
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