Unlock instant, AI-driven research and patent intelligence for your innovation.

Ultra low temperature freezing-storage method for onosma paniculatum cell

A cryopreservation method, the technique of comfrey yunnanensis, applied to plant cells, etc., to achieve the effect of simple operation, good cryopreservation effect, and high cell survival rate

Inactive Publication Date: 2010-01-20
KUNMING UNIV OF SCI & TECH
View PDF3 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The key issues of using liquid nitrogen cryopreservation method are: 1. The selection of protective agent components and the concentration of each component; 2. The operation steps of cryopreservation freezing stage and thawing recovery stage

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0016] 1) Subculture of Comfrey yunnanensis cells: The subculture medium is: MS basal medium + NAA 1.0mg / L + 2, 4-D 0.2mg / L, the culture conditions are solid culture, pH value 5.6, temperature 25 ℃, cultured in the dark;

[0017] 2) Pre-cultivation of Lilacia yunnanensis cells before ultra-low temperature treatment: the cells subcultured for 9 days were transferred to the subculture medium (same as the subculture medium above) containing 5% dimethylsulfoxide (DMSO) for preculture 6 days;

[0018] 3) Dehydration treatment: use 20% glycerol + 10% ethylene glycol + 10% dimethyl sulfoxide + 60% MS basal medium + 0.3mol / L sucrose as components to prepare a cryoprotectant, put the cells in 20 ℃ cryoprotectant for 15 minutes, and then pre-cooled at 0 ℃ for 30 minutes;

[0019] 4) Freezing process: Put the pre-cooled cells and cryoprotectant into liquid nitrogen for freezing;

[0020] 5) Thawing process: Take out the frozen cells from liquid nitrogen, thaw them in a water bath at 2...

Embodiment 2

[0023] 1) Subculture of Comfrey yunnanensis cells: The subculture medium is: MS basal medium + NAA 1.0mg / L + 2, 4-D 0.2mg / L, the culture conditions are solid culture, pH value 5.6, temperature 25 ℃, cultured in the dark;

[0024] 2) Pre-cultivation of B. yunnanensis cells before ultra-low temperature treatment: transfer the cells subcultured for 10 days to the subculture medium (same as the subculture medium above) containing 9% dimethylsulfoxide (DMSO) for preculture 4 days;

[0025] 3) Dehydration treatment: use 25% glycerol + 10% ethylene glycol + 8% dimethyl sulfoxide + 57% MS basal medium + 0.4mol / L sucrose as components to prepare a cryoprotectant, put the cells in 15 ℃ cryoprotectant for 20 minutes, and then pre-cooled at 0 ℃ for 30 minutes;

[0026] 4) Freezing process: Put the pre-cooled cells and cryoprotectant into liquid nitrogen for freezing;

[0027] 5) Thawing process: Take out the frozen cells from liquid nitrogen, thaw them in a water bath at 25°C, and then...

Embodiment 3

[0030] 1) Subculture of Comfrey yunnanensis cells: The subculture medium is: MS basal medium + NAA 1.0mg / L + 2, 4-D 0.2mg / L, the culture conditions are solid culture, pH value 5.6, temperature 25 ℃, cultured in the dark;

[0031] 2) Pre-cultivation of B. yunnanensis cells before ultra-low temperature treatment: the cells subcultured for 11 days were transferred to the subculture medium (same as the subculture medium above) containing 10% dimethylsulfoxide (DMSO) for preculture 8 days;

[0032] 3) Dehydration treatment: use 30% glycerol + 20% ethylene glycol + 5% dimethyl sulfoxide + 45% MS basal medium + 0.6mol / L sucrose as components to prepare a cryoprotectant, put the cells in 25 ℃ cryoprotectant for 30 minutes, and then pre-cooled at 0 ℃ for 30 minutes;

[0033] 4) Freezing process: Put the pre-cooled cells and cryoprotectant into liquid nitrogen for freezing;

[0034] 5) Thawing process: Take out the frozen cells from liquid nitrogen, thaw them in a water bath at 10°C,...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention relates to a super cryopreservation method for onosma paniculatum cells. The method mainly comprises the operating steps: firstly, pretreatment and dehydration process before the cryopreservation of the cells; secondly, cryopreservation stage of the cells in liquid nitrogen; thirdly; thawing process of the cryopreservation cells; and fourthly, resuscitation and culturing of the cells after the thawing. When the onosma paniculatum cells are preserved by utilizing the super cryopreservation method, the cells can obtain higher survival rate after being thawed, and the method is an effective way for preserving the onosma paniculatum germplasm resource.

Description

technical field [0001] The invention relates to a method capable of ultra-low temperature cryopreservation of Lilacia yunnanensis cells. Background technique [0002] Yunnan comfrey (Onosmapaniculatum Bur.et Fr.) is mainly produced in Yunnan. It is a commonly used Chinese herbal medicine in Yunnan Province. It belongs to Boraginaceae (Boraginaceae), and its root is used for medicine. The main active ingredients are shikonin and its derivatives. These ingredients not only have various pharmacological effects such as antibacterial, anti-inflammatory, and anti-cancer, but also are widely used in medicine, cosmetics, and printing and dyeing industries as natural pigments. At present, the wild resources of yunnanensis are severely damaged, and the contradiction between supply and demand in the market is prominent. The use of large-scale plant cell culture technology to directly cultivate cells in reactors to obtain medicinal secondary metabolites is the most direct and effective...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/04
Inventor 葛锋陈朝银韩本勇熊向峰
Owner KUNMING UNIV OF SCI & TECH