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Method for cultivation and production of paclitaxel from induction of taxus chinensis var. mairei axillary bud in vitro

A technology of southern yew and paclitaxel, applied in the biological field, can solve problems such as the proliferation of axillary buds of the southern yew, and achieve the effects of short and thick axillary buds, short induction time, and high induction rate

Inactive Publication Date: 2012-08-15
JISHOU UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although these methods all have good effects on the rapid propagation of Taxus chinensis, they do not mention the use of TIBA to induce the proliferation of Taxus chinensis axillary buds.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] (1) Cut the young shoots of the year from the southern yew trees, remove the leaves, and cut them into small stems of 2.5 cm.

[0022] (2) Add tap water into the beaker, add a small amount of washing powder into the water, add 0.5g of washing powder per 500mL of water, and stir evenly; then put the cut small stems into the beaker and stir for 30 minutes, take it out, and rinse with tap water After 2 hours, transfer to a sterile ultra-clean workbench, soak and disinfect with 70% alcohol for 30 seconds, wash with sterile water for 4 times, and then use 0.1% HgCl 2 The solution was soaked and disinfected for 8 minutes, washed with sterile water for 6 times, and the surface water was dried in an ultra-clean workbench for backup inoculation.

[0023] (3) Insert the morphological lower end of the explant prepared in step (2) with 0.1 mg / L of 2,3,5-triiodobenzoic acid (TIBA for short) Axillary bud induction culture was carried out in WPM medium. The improved WPM medium is ba...

Embodiment 2

[0028] (1) Cut the young shoots of the year from the southern yew trees, remove the leaves, and cut them into small stems of about 2 cm.

[0029] (2) Add tap water into the beaker, add a small amount of washing powder in the water, add 0.5g of washing powder per 500mL of water, and stir evenly; then put the cut small stems into the beaker and stir for 30 minutes, take it out, and rinse with tap water After 2 hours, transfer to a sterile ultra-clean workbench, soak and disinfect with 72% alcohol for 30 seconds, wash with sterile water for 4 times, and then use 0.2% HgCl 2 The solution was soaked and disinfected for 8 minutes, washed with sterile water for 6 times, and the surface water was dried in an ultra-clean workbench for backup inoculation.

[0030] (3) Insert the morphological lower end of the explant prepared in step (2) into the modified WPM supplemented with 3.0 mg / L 2,3,5-triiodobenzoic acid (TIBA for short) Induction culture of axillary buds in culture medium. The...

Embodiment 3

[0036] (1) Cut the young shoots of the year from the southern yew trees, remove the leaves, and cut them into stems of about 3 cm.

[0037] (2) Add tap water into the beaker, add a small amount of washing powder in the water, add 0.5g of washing powder per 500mL of water, and stir evenly; then put the cut stem into the beaker and stir for 30 minutes, take it out, and rinse it with tap water for 2 hours Then transfer to a sterile ultra-clean workbench, soak and disinfect with 75% alcohol for 30 seconds, wash with sterile water for 4 times, and then use 0.25% HgCl 2 The solution was soaked and disinfected for 8 minutes, washed with sterile water for 6 times, and the surface water was dried in an ultra-clean workbench for backup inoculation.

[0038] (3) Insert the morphological lower end of the explant prepared in step (2) into the modification with 4.0mg / L 2,3,5-triiodobenzoic acid (2,3,5-Triiodobenzoic acid, TIBA for short) Axillary bud induction culture was carried out in WPM ...

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PUM

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Abstract

The invention provides a method for extracting paclitaxel from induction of taxus chinensis var. mairei axillary bud germination in vitro and then cultivation and production. On the basis of a WPM (woody plant medium) minimal medium, the method uses (800-1000) mg / L of (NH4)2SO4 to replace 990 mg / L of K2SO4 to obtain the improved WPM minimal medium, and adding 0.01-4.5 mg / L of TIBA; pruning annualyoung shoots from a taxus chinensis var. mairei tree, cutting into small stem sections in 2.5cm, sterilizing, inoculating to the improved WPM minimal medium, and performing alternative culture, namely light culture and dark culture, in an artificial climate box for culturing temperature, wherein the temperature is 25+ / -1 DEG C in the daytime and is 20+ / -1 DEG C in the night, the illumination intensity is 1500lx, and relative humidity is 80%, thus rapidly and successfully inducing axillary buds based on the WPM plus 0.01-4.5 mg / LTIBA. By using the method, the industrialization of the taxus chinensis var. mairei axillary buds can be realized with large scale, short period, high reproduction rate and low cost. Buds can be induced around 15 days with 100% induction rate; and the taxus chinensis var. mairei axillary buds induced by the method is thick and short and has freshly green color. The content of paclitaxel in buds obtained by the method can reach 0.0042% in fifty days, which is higher than that of paclitaxel in natural buds.

Description

technical field [0001] The invention relates to the field of biology, in particular to a method for producing paclitaxel by in vitro induction and culture of axillary buds of Taxus chinensis. Background technique [0002] Yew is recognized as an endangered natural and rare anti-cancer plant in the world. It is an ancient tree species left over from Quaternary glaciers with a history of 2.5 million years. The yew plant is distributed in the northern hemisphere. It is a tall tree with a sparse population distribution, slow growth rate and poor regeneration ability. Therefore, for a long time, there has not been a large-scale yew raw material forest base in the world. my country has listed it as a first-class rare and endangered protected plant, and the United Nations has also banned logging. There are eleven species of yew in the world, the southern yew ( Taxus chinensis var. mirei ) is a variety of yew in China. [0003] Amos et al. (1981) found that adding low concentr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01H4/00
Inventor 杜亚填张翔宇龚雪原
Owner JISHOU UNIVERSITY
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