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Rubber tree embryogenic callus induction medium and rubber tree embryogenic callus rapid proliferation method

A technology of embryogenic callus and induction medium, applied in horticultural methods, botanical equipment and methods, plant regeneration, etc., can solve the problem of long callus cycle, slow speed of sterile seedlings, and easy degeneration of sterile seedlings Or mutation and other issues, to promote differentiation and division, avoid content decline, and enhance the effect of division

Active Publication Date: 2019-03-12
HAINAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Obtaining regenerated plants through organogenesis and rapid propagation has become a more effective propagation method, but there is still a long cycle of explant-induced callus, slow speed of obtaining sterile seedlings, and long-term continuous proliferation of sterile seedlings Very prone to degeneration or mutation and other shortcomings

Method used

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  • Rubber tree embryogenic callus induction medium and rubber tree embryogenic callus rapid proliferation method
  • Rubber tree embryogenic callus induction medium and rubber tree embryogenic callus rapid proliferation method
  • Rubber tree embryogenic callus induction medium and rubber tree embryogenic callus rapid proliferation method

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0074] The material is Hevea brasiliensis Müll.Arg. variety Reyan 7-33-97, and the rubber inflorescences were collected from the experimental farm of the Chinese Academy of Tropical Agricultural Sciences. Submerge the inflorescences with 0.1% light soapy water ( figure 1 -A) 10min, rinse with running water. Use sterilized small scissors to cut out unopened, healthy rubber tree male flowers ( figure 1 -B), most of the pollen grain development period is the uninucleate marginal stage ( figure 1 -D). Wrap it with sterile gauze, disinfect the surface with 70-75% alcohol for 40 seconds, and use 0.1% HgCl 2 Soak and disinfect for 12 minutes, then rinse with sterile water for 6 times, and then peel off a single anther under a dissecting microscope and inoculate it on the anther callus induction medium, with 30 anthers per dish.

[0075] Table 1 Correspondence between the pollen development period and the external shape of flower buds of rubber tree Reyan 7-33-97 variety

[0076]...

Embodiment 2

[0090] Somatic embryonic differentiation of embryogenic callus

[0091] The selected embryogenic calli with spherical particles on the surface were connected to the somatic embryo induction medium, subcultured once every 25 days, and cultured in the dark at a temperature of 25°C during the day, 23°C at night, and a humidity of 80%. After 50 days, the embryogenic callus gradually differentiated into somatic embryoid bodies at different developmental stages, accompanied by the proliferation of embryogenic callus. Somatic embryo differentiation medium is based on modified MS, supplemented with Kt 0.6mg / L, NAA 0.15mg / L, 6-BA 0.05mg / L, spermidine 5mg / L, hydrolyzed casein 300mg / L, 10% Coconut milk, active carbon 0.10%, sucrose 70g / L and plant gel 2.4g / L, the pH value of culture medium is 5.8. The improved basic culture of MS is: KH 2 PO 4 226mg / L, NH 4 NO 3 1100mg / L, KNO 3 1267mg / L, CaCl 2 2H 2 O800mg / L, MgSO 4 ·7H 2 O 247mg / L, H 3 BO 3 9mg / L, in organic ingredients, thi...

Embodiment 3

[0096] Rapid proliferation of embryogenic callus

[0097]With the help of a dissecting microscope, undifferentiated embryogenic calli with compact texture and spherical granules on the surface and normal cotyledon-shaped somatic embryoids (single cotyledons about 2-3mm wide), inoculated together on the proliferation medium, nursed and nurtured by normal cotyledon-shaped somatic embryoid bodies, so that the embryogenic callus can proliferate effectively and rapidly. Cultivate in indoor natural scattered light for 25-30 days, the temperature is 25±2°C during the day, 23±2°C at night, and the humidity is 80%. The embryoid body germination medium uses the improved MS as the basic medium, adds hydrolyzed casein 300mg / L, 10% coconut milk, sucrose 70g / L, 0.1% activated carbon and plant gel 2.3g / L, pH5.8, without adding any hormones.

[0098] The above-mentioned undifferentiated embryogenic callus with compact texture and spherical granules on the surface was inserted into the proli...

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Abstract

The invention provides an induction culture medium for embryogenic callus of hevea brasiliensis and a rapid propagation method for embryogenic callus of hevea brasiliensis, and belongs to the technical field of tissue culture. The induction culture medium is an improved MS culture medium containing 2, 4-D-compounded 6-BA, zeatin, TDZ, coconut milk and sucrose, and greatly increases an induction rate of embryogenic callus. The rapid propagation method for embryogenic callus of hevea brasiliensis comprises the following steps: performing anther primary induction, embryogenic callus induction and somatic embryoid differentiation culture to respectively obtain embryogenic callus and cotyledon-like somatic embryoid; simultaneously inoculating the embryogenic callus and the cotyledon-like somatic embryoid into a proliferation culture medium; and using the cotyledon-like somatic embryoid for nurse breeding, so that the embryogenic callus is proliferated effectively and rapidly. With germination of cotyledon-like somatic embryos, the embryogenic callus is rapidly proliferated, so that the cotyledon-like somatic embryos and the embryogenic callus have good synchronicity and are both in an embryogenic undifferentiated state of the embryogenic callus.

Description

technical field [0001] The invention belongs to the technical field of tissue culture, in particular to a rubber tree embryogenic callus induction medium and a rubber tree embryogenic callus rapid proliferation method. Background technique [0002] Hevea brasiliensis Müll. Arg. belongs to Euphorbiaceae (Euphorbiaceae) rubber tree genus (Hevea), is a perennial cross-pollinated tree. Rubber tree is the main source of natural rubber, while the self-sufficiency rate of natural rubber in my country is only 18%. At present, my country's natural rubber import sources are highly concentrated in Southeast Asian countries such as Thailand, Indonesia, Malaysia, and Vietnam. The high concentration of import sources, geopolitical instability and fierce competition for resources have increased the uncertainty and potential risks of my country's natural rubber imports. In view of the limited area of ​​rubber planting areas in my country, increasing the output of rubber trees is an effect...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A01H4/00
CPCA01H4/001A01H4/008
Inventor 陈健妙何朝族周广振
Owner HAINAN UNIV
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