Method for obtaining callus tissue of stevia rebaudiana bertoni

A callus and callus induction technology, applied in the field of stevia callus acquisition, can solve the problems of loss of division ability and no reports, and achieve the advantages of short establishment period, unique technology, and shortened selection and cultivation time. Effect

Inactive Publication Date: 2018-05-04
BEIJING AGRO BIOTECH RES CENT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, guard cells have long been recognized as terminally differentiated specialized cells that have lost the ability to divide
At present, there are no reports at home and abroad on the isolation and in vitro culture of stomatal guard cells in Stevia rebaudiana leaves

Method used

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  • Method for obtaining callus tissue of stevia rebaudiana bertoni
  • Method for obtaining callus tissue of stevia rebaudiana bertoni
  • Method for obtaining callus tissue of stevia rebaudiana bertoni

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0097] This example is used to illustrate the method for obtaining the Stevia rebaudiana callus of the present invention.

[0098] (1) Acquisition of epidermal fragments (containing stomatal guard cells) of Stevia rebaudiana

[0099] Stevia epidermal fragments (containing stomatal guard cells) were obtained as follows:

[0100] Stevia rebaudiana seeds are cultured and germinated in a cultivation medium to obtain stevia rebaudiana sterile seedlings. Select robust stevia aseptic seedling plants with vigorous growth, thick stems and thick leaves, large and flat leaves, and bright green leaves, and transfer them to a new cultivation medium for the subculture of the first subculture cycle (28 days) , then transfer to a new cultivation medium to continue the subculture of the second subculture cycle (28 days), then transfer to a new cultivation medium to continue the third subculture cycle (28 days) ) subculture, select the stevia tissue culture sterile plant that adopts in the pr...

Embodiment 2

[0127] This example is used to illustrate the method for obtaining the Stevia rebaudiana callus of the present invention.

[0128] Except the following operating parameters, all the other are the same as Example 1.

[0129] (1) Acquisition of epidermal fragments (containing stomatal guard cells) of Stevia rebaudiana

[0130] The skimmer operates for 1 second in "low speed" mode and for 30 seconds in "high speed" mode.

[0131] (2) Induction culture

[0132] The induction medium is: based on the B5 basic medium, adding sucrose to a final weight percentage of 3%, adding 6-BA to a final concentration of 0.5 mg / L, adding NAA to a final concentration of 0.5 mg / L, Glutamine was added to a final concentration of 100 mg / L.

[0133] The division rate of stomatal guard cells can reach 3.0%; the number of cells contained in the obtained small cell mass is 10; the rate of planting plate is 0.15%.

[0134] (3) Suspension culture

[0135] After the induced culture product (containing s...

Embodiment 3

[0142] This example is used to illustrate the method for obtaining the Stevia rebaudiana callus of the present invention.

[0143] Except the following operating parameters, all the other are the same as Example 1.

[0144] (1) Acquisition of epidermal fragments (containing stomatal guard cells) of Stevia rebaudiana

[0145] The skimmer operates for 1 second in "low speed" mode and for 30 seconds in "high speed" mode.

[0146] (2) Induction culture

[0147] The induction medium is: based on the B5 basic medium, adding sucrose to a final weight percentage of 3%, adding 6-BA to a final concentration of 5.0 mg / L, adding NAA to a final concentration of 0.05 mg / L, Add glutamine to a final concentration of 200 mg / L.

[0148] The first addition of the liquid medium: the volume ratio of the liquid medium to the original medium is 0.5:10;

[0149] Add liquid medium for the second time: the volume ratio of liquid medium and original medium is 1:10.

[0150] The division rate of sto...

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Abstract

The invention belongs to the field of plant cell culture, and particularly relates to a method for obtaining callus tissue of stevia rebaudiana bertoni. The method for obtaining the callus tissue of the stevia rebaudiana bertoni comprises steps as follows: induced culture of stomatal guard cells of the stevia rebaudiana bertoni, suspension culture and induced culture of the callus tissue. With theadoption of the method for obtaining the callus tissue of the stevia rebaudiana bertoni, a large quantity of epithelial fragments containing the stomatal guard cells can be quickly obtained, a high-purity suspension cell culture system is rapidly established, cell screening and culture time is shortened, the obtained callus tissue is high in quality, the percentage of embryonic callus is high, and weight increment is fast.

Description

technical field [0001] The invention belongs to the field of plant cell culture, and in particular relates to a method for obtaining stevia callus. Background technique [0002] Stevia (Stevia rebaudiana Bertoni) is a plant of the genus Stevia in the family Asteraceae, native to Paraguay in South America, and introduced to China from Japan in 1976. Now, my country has become the country with the largest planting area of ​​stevia in the world, and it is planted in large areas in Jiangsu, Fujian, Guangdong, Zhejiang, Shandong, Henan, Anhui, Xinjiang and other places. Stevia leaves are rich in stevia glycosides, high in sweetness (350-400 times that of sucrose), low in calories (1 / 300 of sucrose), safe and non-toxic, and known as "the third healthy sugar source in the world". In 1985 and 1990, the Chinese Ministry of Health approved steviol glycosides as natural sweeteners and pharmaceutical sweetener excipients for unlimited use. In addition, steviol glycosides can control b...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01H4/00
CPCA01H4/005
Inventor 李宏潮张秀海陈绪清杜运鹏
Owner BEIJING AGRO BIOTECH RES CENT
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