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Vincaleukoblastinum-rich catharanthus roseus cell culturing method

A cell culture and periwinkle technology, applied in the field of cell culture, can solve the problems of high production cost of vinblastine and high price of vinblastine, and achieve the effects of high accumulative content of vinblastine, shortening induction period and avoiding browning.

Inactive Publication Date: 2019-03-12
覃家日
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The content of vinblastine in vinblastine cells is relatively low, resulting in higher production costs per unit mass of vinblastine. Even if vinblastine is produced through cell culture, the price of vinblastine is still relatively high, which cannot meet market demand

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] A method for culturing vinblastine cells rich in vinblastine, comprising the following steps:

[0022] 1) Select plump periwinkle seeds, soak them in weak acid electrolyzed water for 10 seconds, wash them with sterile deionized water, and use them on the B5 foundation containing 10mg / L vitamin C, 7g / L agar and 0.1g / L oligosaccharin The aseptic seedlings were cultured on the culture medium, and the hypocotyls of the aseptic seedlings were used as explants, which were inserted into the callus induction medium for induction culture. The culture temperature was 32°C, and the light and dark cultures were alternately carried out every day. Induce the callus, then transfer the callus to the subculture medium, subculture once every 25 days, and subculture 3 times to obtain stable periwinkle callus;

[0023] 2) Select the periwinkle callus with vigorous growth, loose texture, and stable and uniform state, inoculate it into a liquid suspension medium, then add glass beads to the ...

Embodiment 2

[0027] A method for culturing vinblastine cells rich in vinblastine, comprising the following steps:

[0028] 1) Select plump periwinkle seeds, soak them in weak acid electrolyzed water for 10 seconds, wash them with sterile deionized water, and use them on the B5 foundation containing 10mg / L vitamin C, 7g / L agar and 0.1g / L oligosaccharin The aseptic seedlings were cultured on the culture medium, and the hypocotyls of the aseptic seedlings were used as explants, which were inserted into the callus induction medium for induction culture. The culture temperature was 32°C, and the light and dark cultures were alternately carried out every day. Induce the callus, then transfer the callus to the subculture medium, subculture once every 25 days, and subculture 3 times to obtain stable periwinkle callus; wherein, the callus induction Medium is the B5 basal medium containing 1mg / L cytokinin, 0.5mg / L 6-benzylaminopurine, 30g / L sucrose, 3mg / L licorice flavone and 8g / L agar;

[0029] 2)...

Embodiment 3

[0033] A method for culturing vinblastine cells rich in vinblastine, comprising the following steps:

[0034] 1) Select plump periwinkle seeds, soak them in weak acid electrolyzed water for 10 seconds, wash them with sterile deionized water, and use them on the B5 foundation containing 10mg / L vitamin C, 7g / L agar and 0.1g / L oligosaccharin The aseptic seedlings were cultured on the culture medium, and the hypocotyls of the aseptic seedlings were used as explants, which were inserted into the callus induction medium for induction culture. The culture temperature was 32°C, and the light and dark cultures were alternately carried out every day. Induce the callus, then transfer the callus to the subculture medium, subculture once every 25 days, and subculture 3 times to obtain stable periwinkle callus; wherein, the callus induction Medium is the B5 basal medium containing 1mg / L cytokinin, 0.5mg / L 6-benzylaminopurine, 30g / L sucrose, 3mg / L licorice flavonoids and 8g / L agar; Add phen...

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PUM

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Abstract

The invention provides a vincaleukoblastinum-rich catharanthus roseus cell culturing method. The vincaleukoblastinum-rich catharanthus roseus cell culturing method comprises the following steps: (1) culturing catharanthus roseus calli; (2) culturing a steadily hereditary catharanthus roseus cell line; (3) inoculating the catharanthus roseus cell line which is obtained in the step (2) into a liquidsuspension culturing medium for performing induced culturing to stimulate the accumulation of alkaloids in catharanthus roseus, wherein the induction conditions are that: from the tenth to fifteenthdays of culturing the catharanthus roseus cell line in the liquid suspension culturing medium, carbon monoxide gas is introduced, and nicotinic acid, ecdysone and silver thiosulfate are added into theliquid suspension culturing medium, and irradiation is performed by using the ultraviolet light with the wavelength of 360 to 400 nm; the irradiation intensity is 10 to 20 mu W / cm<2>; the irradiationis stopped for 30 to 45 minutes after the irradiation is performed for 10 to 15 minutes until the culturing is ended; and vincaleukoblastinum-rich catharanthus roseus cells are obtained. The method has the characteristics of high cell growth speed, high vincaleukoblastinum accumulation content and the like.

Description

technical field [0001] The invention belongs to the technical field of cell culture. More specifically, the present invention relates to a method for culturing vinblastine-rich vinca cells. Background technique [0002] Periwinkle, also known as Calendula, Sishichun, Ririxin, Yantouhong, Sanwanhua, is a plant of the genus Periwinkle in the family Apocynaceae. At present, more than 100 kinds of terpene indole alkaloids have been isolated from Vinca, such as Vinblastine, Vincristine, Serpentine, Wendoline, etc. Many alkaloids isolated from Vinca have Antitumor activity, among them, vinblastine is the alkaloid with the greatest medicinal value and the most widely used in Changchun flower. The content of vinblastine in vinblastine cells is low, resulting in higher production costs per unit mass of vinblastine. Even if vinblastine is produced through cell culture, the price of vinblastine is still relatively high, which cannot meet market demand. Contents of the invention ...

Claims

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Application Information

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IPC IPC(8): C12N5/04C12P17/18
CPCC12N5/04C12P17/182
Inventor 覃家日
Owner 覃家日
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