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Method for inducing rhizoma bletillae tetraploid through calluses

A callus induction and callus technology, applied in the field of plant biology, can solve problems such as test failure, pollution, and increased seed contamination rate

Active Publication Date: 2016-06-08
广西壮族自治区农业科学院生物技术研究所
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, the use of chemical inducers to induce polyploidy in the seeds of Bletilla has been reported. Because the seeds of Bletilla are small, when washing after induction, the seeds are suspended in water. Although it can be solved by centrifugation, because the seeds of Bletilla must be It can only germinate on the culture medium, and repeated cleaning work between the ultra-clean workbench and the centrifuge will greatly increase the seed contamination rate, and in severe cases, the test will fail due to the contamination of the entire treatment

Method used

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  • Method for inducing rhizoma bletillae tetraploid through calluses

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] A method for inducing white and tetraploidy with callus, comprising the following steps:

[0029] (1) On the ultra-clean workbench, cut the callus into small pieces of 0.3cm×0.3cm, insert them into the proliferation medium and cultivate them for 10 days; under sterile conditions, add sterilized chemical Inducer, chemical inducer soaked callus for 24h;

[0030] The chemical inducer is: 0.05% colchicine+3%DMSO;

[0031] The proliferation medium is: N 6 +2,4-D1.0mg / L+NAA0.5mg / L+TDZ0.5mg / L, pH6.0;

[0032] (2) On the ultra-clean workbench, take out the callus in step (1), wash it in sterile water for 5 times, blot the water on sterile filter paper, insert it into the differentiation medium and culture it, and produce Bud point, 36d to form a bud, 55d to grow 2-3 leaves, the bud is thin and weak, and the leaf color is yellow-green;

[0033] The differentiation medium is: MS+6-BA1.0mg / L+ZT0.5mg / L+NAA0.5mg / L, pH6.0;

[0034] (3) On the ultra-clean workbench, put the buds ...

Embodiment 2

[0041] (1) On the ultra-clean workbench, cut the callus into small pieces of 0.3cm×0.3cm, insert them into the proliferation medium and cultivate them for 10 days; under sterile conditions, add sterilized chemical Inducer, chemical inducer soaked callus for 8h;

[0042] The chemical inducer is: 0.2% colchicine+3%DMSO;

[0043] The proliferation medium is: N 6 +2,4-D1.0mg / L+NAA0.5mg / L+TDZ0.5mg / L, pH6.0;

[0044] (2) On the ultra-clean workbench, take out the callus in step (1), wash it in sterile water for 5 times, blot the water with sterile filter paper, insert it into the differentiation medium for culture, and partially callus The tissue turns black, and it takes a long time to induce budding. Bud points will appear in 30 days, buds will form in 43 days, and 2-3 leaves will grow in 65 days. The buds are thin and thick, and the leaves are yellow-green and dark green;

[0045] The differentiation medium is: MS+6-BA1.0mg / L+ZT0.5mg / L+NAA0.5mg / L, pH6.0;

[0046] (3) On the u...

Embodiment 3

[0053] (1) On the ultra-clean workbench, cut the callus into small pieces of 0.3cm×0.3cm, insert them into the proliferation medium and cultivate them for 10 days; under sterile conditions, add sterilized chemical Inducer, chemical inducer soaked callus for 16h;

[0054] The chemical inducer is: 0.1% colchicine+3%DMSO;

[0055] The proliferation medium is: N 6 +2,4-D1.0mg / L+NAA0.5mg / L+TDZ0.5mg / L, pH6.0;

[0056] (2) On the ultra-clean workbench, take out the callus in step (1), wash it in sterile water for 5 times, blot the water with sterile filter paper, insert it into the differentiation medium for culture, and produce buds after 25 days 40d to form buds, 60d buds grow 2-3 leaves, the buds are thick, and the leaves are dark green;

[0057] The differentiation medium is: MS+6-BA1.0mg / L+ZT0.5mg / L+NAA0.5mg / L, pH6.0;

[0058] (3) Put the buds obtained in step (2) into the rooting medium on the ultra-clean workbench and cultivate them. On the 7th day, the lower end of the bu...

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Abstract

The invention discloses a method for inducing rhizoma bletillae tetraploid through calluses. A chemical inducer comprising 0.05%-0.2% of colchicines and 3% of DMSO is used for conducting short-time induction on the calluses, proliferation, differentiation and rooting culture are conducted, and the inductivity of the rhizoma bletillae tetraploid is 2.6%-23.4% through experimental statistics. According to the method, the inductivity of the tetraploid can be increased, and the toxic effect of the chemical inducer on cells can be relieved. Based on the cell totipotency principle, in the differentiation process, one cell is differentiated into a plant, and heterozygotes can be effectively reduced. Thus, the method has the advantages that operation is simple, control is easy, repeatability is good, inductivity is high, and the purification rate of polyploidy is high.

Description

technical field [0001] The invention belongs to the field of plant biotechnology, and in particular relates to a method for inducing tetraploids with callus tissue. Background technique [0002] White and ( Bletilla striata (Thunb.) Reichb.f.) also known as Bletilla striata, alias purple orchid, is a perennial herbaceous plant in the family Orchidaceae. Its tuber is a traditional Chinese medicine in my country. It is slightly cold in nature, bitter in taste and sweet in taste. ; It is an important component of Chinese patent medicines such as "Kaiwei Tablet" and "Weikangning". In addition, Baiji also has important application value in beauty and skin care, fine chemicals and ornamental gardening. [0003] At present, white and raw materials mainly rely on wild natural resources. Excessive collection and habitat destruction have drastically reduced the number of resources and are on the verge of extinction. With the continuous deepening of research on white and its wide ap...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01H4/00
CPCA01H4/005
Inventor 石云平苏祖祥韦绍龙林茜李小泉
Owner 广西壮族自治区农业科学院生物技术研究所
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