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Inducing method and proliferation method of red clover hairy roots

A technology of red clover and hairy roots is applied in the field of induction and proliferation of red clover hairy roots to achieve the effects of rapid growth, improved induction transformation rate and survival rate, and improved growth cycle.

Inactive Publication Date: 2020-04-10
INNER MONGOLIA UNIV FOR THE NATITIES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] In addition, although in the prior art, Agrobacterium rhizogenes can infect most dicotyledonous plants and a few monocotyledonous plants, and can induce hairy roots by infecting plants with Agrobacterium rhizogenes; however, the prior art has not been successful Efficient induction of red clover by Agrobacterium rhizogenes and systematic optimization of the liquid culture system of the hairy root system

Method used

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  • Inducing method and proliferation method of red clover hairy roots
  • Inducing method and proliferation method of red clover hairy roots
  • Inducing method and proliferation method of red clover hairy roots

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Embodiment 1

[0035] This embodiment provides a kind of induction method of red clover hairy root, it comprises the following steps:

[0036] (1) After the red clover seeds are disinfected and cleaned, the red clover seeds are placed in MS liquid medium for cultivation to obtain red clover aseptic seedlings, as attached figure 1 As shown; specifically, take plump red clover seeds first, soak them in warm water for 1 hour, and then wash them with sterile water for 3 times; then, carry out preliminary disinfection of red clover seeds with 90% ethanol solution for 40 seconds Finally, the red clover seeds after preliminary disinfection are cleaned 3 times with sterile water, each time for 30s; After disinfection for 7 minutes, the red clover seeds were washed 10 times with sterile water, each time for 3 minutes; then, the washed red clover seeds were placed on filter paper to germinate, and then placed in MS liquid medium (Guarantee 100% germination of seeds and robustness of seedlings), and p...

Embodiment 2

[0041] This embodiment provides a kind of induction method of red clover hairy root, it comprises the following steps:

[0042] (1) After the red clover seeds are disinfected and cleaned, then the red clover seeds are placed in MS liquid medium for cultivation to obtain red clover aseptic seedlings; specifically, first get full red clover seeds, After soaking in warm water for 1 hour, wash with sterile water for 3 times; then, use 75% ethanol solution to preliminarily sterilize the red clover seeds for 40 seconds, and then use sterile water to clean the red clover seeds Bacteria water was cleaned 3 times, and each cleaning was 30s; Then, the red clover seeds after cleaning were sterilized again with 0.1% mercuric chloride solution for 7 minutes, and then the red clover seeds were treated with sterile water. Carry out cleaning 10 times, each cleaning 3min; Then, after the red clover seeds after cleaning are placed on the filter paper to germinate, then be placed in MS liquid cu...

Embodiment 3

[0047] This embodiment provides a kind of induction method of red clover hairy root, it comprises the following steps:

[0048] (1) After the red clover seeds are disinfected and cleaned, then the red clover seeds are placed in MS liquid medium for cultivation to obtain red clover aseptic seedlings; specifically, first get full red clover seeds, After soaking in warm water for 1 hour, wash with sterile water for 3 times; then, use 75% ethanol solution to preliminarily sterilize the red clover seeds for 40 seconds, and then use sterile water to clean the red clover seeds Bacteria water was cleaned 3 times, and each cleaning was 30s; Then, the red clover seeds after cleaning were sterilized again with 0.1% mercuric chloride solution for 7 minutes, and then the red clover seeds were treated with sterile water. Carry out cleaning 10 times, each cleaning 3min; Then, after the red clover seeds after cleaning are placed on the filter paper to germinate, then be placed in MS liquid cu...

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Abstract

The invention discloses an inducing method and proliferation method of red clover hairy roots, and belongs to the technical field of biology. The inducing method comprises the following steps of firstly, performing disinfection and cleaning treatment on red clover seeds, placing the treated red clover seeds in MS liquid culture mediums, and performing culture so as to obtain red clover bacteria-free seedlings; with the red clover bacteria-free seedlings as infestation materials, and injecting agrobacterium rhizogenes infestation liquid into the infestation materials; and then inoculating the MS culture mediums containing acetosyringone, with the infestation materials to which the agrobacterium rhizogenes infestation liquid is injected, performing co-culture treatment, then transferring theco-cultured MS culture mediums into MS culture mediums containing cefotaxime sodium, and performing bacteriostasis culture treatment so as to obtain induced converted hairy roots. According to the inducing method disclosed by the invention, through screening appropriate explants and improving a bacterial liquid infestation manner, besides, through controlling the addition concentration of the acetosyringone and the cefotaxime sodium and controlling co-culture time, the inducing conversion rate and the survival rate of the hairy roots can be greatly increased.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for inducing and multiplying hairy roots of red clover. Background technique [0002] Red clover isoflavones are important secondary metabolic active ingredients, which have a variety of important pharmacological effects. [0003] Has high medical value. Although the cultivated red clover is rich in resources, its growth cycle is long, and the content of active components is easily affected by factors such as environmental changes and pests and diseases, which may easily cause the problem of decline in the yield and quality of medicinal materials. In contrast, the hairy root culture system has the characteristics of genetic stability, rapid growth, safety and reliability, high synthesis ability of isoflavone secondary metabolites and large-scale production. Therefore, there is an urgent need to study a method for the induction of red clover hairy roots and the system optim...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01H4/00
CPCA01H4/001A01H4/008
Inventor 于丽丽银铃狄建军王华张学富孙立杰穆莎茉莉
Owner INNER MONGOLIA UNIV FOR THE NATITIES
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