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Method for Tissue Culture Regeneration of Mature Bromegrass Seeds

A technique for tissue culture of brome, applied in the field of agricultural biology, can solve problems such as blanks, and achieve the effects of low cost, simple culture process and large number of plants

Inactive Publication Date: 2016-07-27
GRASSLAND RES INST OF CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the domestic research on the establishment of the in vitro tissue culture regeneration system of brome awnless is still in its infancy. Recently, there has been no report on the establishment of a tissue culture regeneration system for mature brome awned seeds, and the research on the establishment of key steps such as induction, differentiation, and rooting in the regeneration process is still blank.

Method used

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  • Method for Tissue Culture Regeneration of Mature Bromegrass Seeds

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0016] Example 1: figure 1 As shown, the method for tissue culture regeneration of mature brome awnless seeds adopts the following specific process steps.

[0017] (1) Sterilization treatment of explants: select full-bodied brome seeds, shell them and place them in a 50ml triangular flask. Disinfect the surface with 75% ethanol by volume on the ultra-clean workbench for 1 min, rinse with sterile water once; then further disinfect with 0.1% mercuric chloride for 15 min, shaking continuously during this period, and finally rinse with sterile water Rinse 5 times, soak in sterile water at room temperature for 12 hours, let the seeds fully imbibition.

[0018] (2) Induction of callus: Inoculate sterilized brome awn seeds on callus induction medium for culture. The culture conditions are: temperature 26°C, light time 12h / d, light intensity 1500Lux; induction medium: add 1mg / L 2,4-D, 0.5mg / L 6-BA, 25g / L to MS medium L of sucrose and 7g / L of agar, pH 5.8, induction time 30d.

[00...

Embodiment 2

[0025] Embodiment 2: The method for tissue culture regeneration of mature brome awnless seeds adopts the following specific process steps.

[0026] (1) Sterilization treatment of explants: same as in Example 1.

[0027] (2) Callus induction: culture conditions: temperature 25°C, light time 13h / d, light intensity 2000Lux; induction medium: MS+0.8mg / L2,4-D+1mg / L6-BA+25g / L sucrose+7g / L agar, pH value 6.0, induction time 35d;

[0028] (3) Transfer the callus to subculture medium 1 for culture: culture conditions: temperature 27°C, light time 13h / d, light intensity 2000Lux; subculture medium 1: MS+1.5mg / L2,4 -D+0.5mg / L6-BA+25g / L sucrose+7g / L agar, pH value 5.9, culture time 35d.

[0029] (4) Transfer the callus to the differentiation medium for differentiation culture: the culture conditions are: temperature 25°C, light time 15h / d, light intensity 2000Lux; differentiation medium: MS+1.5mg / LNAA+0.8mg / L6- BA+30g / L sucrose+7g / L agar, pH value 6.0, culture time 32d.

[0030] (5) I...

Embodiment 3

[0033] Embodiment 3: The method for tissue culture regeneration of mature brome awnless seeds adopts the following specific process steps.

[0034] (1) Sterilization of explants: same as in Example 1.

[0035] (2) Callus induction: culture conditions: temperature 27°C, light time 14h / d, light intensity 1000Lux; induction medium: MS+0.5mg / L2,4-D+0.7mg / L6-BA+25g / L sucrose +7g / L agar, pH value 5.9, induction time 33d;

[0036] (3) Transfer the callus to subculture medium 1 for culture: culture conditions: temperature 25°C, light time 14h / d, light intensity 1000Lux; subculture medium 1: MS+2mg / L2, 4-D+0 .8mg / L6-BA+25g / L sucrose+7g / L agar, pH value 5.9, culture time 32d.

[0037] (4) Transfer the callus to the differentiation medium for differentiation culture: the culture conditions are: temperature 27°C, light time 16h / d, light intensity 1000Lux; differentiation medium: MS+1.8mg / LNAA+0.5mg / L6- BA+30g / L sucrose+7g / L agar, pH value 5.9, culture time 35d.

[0038] (5) Insert the...

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Abstract

The invention discloses a method for tissue culture and regeneration of mature seeds of awnless brome. The mature seeds of awnless brome are used as explants, and the surface is first sterilized, and then the sterilized seeds of awnless brome are transferred to cultured in the callus induction medium; the induced callus was transferred to the first subculture medium for the first subculture, and then transferred to the differentiation medium for culture; the adventitious buds obtained from the differentiation culture were inserted into the second The second subculture was carried out in the subculture medium, and after differentiation and emergence, the seedlings were inserted into the rooting medium to induce rooting. In this method, mature brome seeds are used as explants, and callus is induced, differentiated and rooted by adding different concentrations of auxin and cytokinin in the medium, so as to establish its complete tissue culture regeneration system. Wheat provides the basis for genetic improvement through genetic engineering.

Description

technical field [0001] The invention belongs to the field of agricultural biotechnology, in particular to a method for culturing and regenerating mature brome seed tissue. Background technique [0002] Bromus inermis is a perennial herbaceous plant belonging to the genus Bromus in the grass family (Gramineae). It has the characteristics of strong adaptability, high nutritional value, large yield, long utilization season, and strong regeneration. It is an excellent forage grass. Its wild species are widely distributed in various provinces and autonomous regions of my country, and it is the dominant grass species in the 1000-3500m mountain meadow. Because of its well-developed underground rhizomes and strong drought resistance, it is also a pioneer plant for establishing artificial pastures and environmental sand fixation. [0003] In recent years, with the adjustment of the industrial structure and the demand for environmental governance, the artificial planting area of ​​b...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A01H4/00
Inventor 黄帆李志勇李鸿雁师文贵刘磊解永凤
Owner GRASSLAND RES INST OF CHINESE ACAD OF AGRI SCI
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