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Method for separating plant endophytes

A plant endophyte and separation method technology, applied in the field of microorganism separation, can solve the problems of limited quantity and type of endophyte, unable to fully represent the quantity and type, etc.

Inactive Publication Date: 2017-08-08
KUNMING UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The technical problem to be solved by the present invention is to overcome the defect that the number and types of endophytic bacteria isolated and obtained are very limited, and the result cannot fully represent the number and types of endophytic bacteria in the whole plant, and provide a kind of plant endophytic bacteria Separation method

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0015] A method for isolating plant endophytes, comprising the following steps:

[0016] 1) Collect healthy, pest-free plant materials, rinse them with tap water, and then wash them with sterile water;

[0017] 2), after disinfecting the surface of the collected material, rinse it with sterile water, dry the surface moisture and set it aside; the disinfection method is to disinfect the collected material in 70% ethanol for 1 min, wash it with sterile water, and then use the volume concentration Soak in 0.1% mercuric chloride for 1 min or soak in 6% sodium hypochlorite for 5 min;

[0018] 3) Cut the sterilized plant materials into small pieces with sterile tools, put them in a container filled with sterile deionized water, and place the sealed container alternately at 4°C and 30°C for 4 to 10 hours;

[0019] 4) Take out the plant material, remove the epidermis, cut into small pieces with sterile tools, grind or add a certain amount of quartz sand in an ice bath to grind into a...

Embodiment 2

[0022] A method for isolating plant endophytes, comprising the following steps:

[0023] 1) Collect healthy, pest-free plant materials, rinse them with tap water, and then wash them with sterile water;

[0024] 2) Rinse the surface of the collected materials with sterile water after disinfection, and dry the surface moisture for later use; the method of disinfection is to sterilize the collected materials with 75% ethanol for 0.5 min, wash them with sterile water, and then use volumetric Soak in 0.3% mercuric chloride for 0.5 min or soak in 4% sodium hypochlorite for 10 min;

[0025] 3) Cut the sterilized plant materials into small pieces with sterile tools, put them in a container filled with sterile deionized water, and put the sealed container in an ice bath at 0°C for 20 minutes, then put it in a water bath at 25-30°C 40min, cycle 4 to 8 times;

[0026] 4) Take out the plant material and remove the epidermis, cut it into small pieces with sterile tools, grind or add a ce...

Embodiment 3

[0029] A method for isolating plant endophytes, comprising the following steps:

[0030] 1) Collect healthy, pest-free plant materials, rinse them with tap water, and then wash them with sterile water;

[0031] 2) Rinse the surface of the collected material with sterile water after disinfection, and dry the surface moisture for later use; the disinfection method is to sterilize the collected material in 72% ethanol for 1 min, wash it with sterile water, and then use the volume concentration Soak in 0.2% mercury chloride for 0.5min or soak in 5% sodium hypochlorite for 6min;

[0032] 3) Cut the sterilized plant materials into small pieces with sterile tools, put them in a container filled with sterile deionized water, place the sealed container at 2°C for 25 minutes, and then put it at 28°C for 35 minutes , cycle 6 times;

[0033] 4) Take out the plant material and remove the epidermis, cut it into small pieces with sterile tools, grind or add a certain amount of quartz sand ...

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PUM

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Abstract

The invention discloses a method for separating plant endophytes. The method comprises the following steps: 1) collecting plant materials, and washing with sterile water; 2) disinfecting the surface and flushing with the sterile water; 3) shearing the materials into small blocks, placing in a container filled with sterile deionized water, sealing the container, alternatively placing the container in an environment of 0-4 DEG C and an environment of 25-30 DEG C for 4-10 hours, and performing endophyte leaching; 4) grinding into homogenate; 5) performing gradient dilution, coating, and transferring single colony subjected to pure culture into test tube slant for preservation. According to the method for separating plant endophytes disclosed by the invention, mixing of infectious microbes is effectively avoided by virtue of disinfection so as to avoid influence of the endophyte separation effect; the plant materials are sealed in the container and alternatively arranged in the environment of 0-4 DEG C and the environment of 25-30 DEG C for 4-10 hours, endophyte leaching is performed, and the plant endophytes are fully released, so that the separated plant endophytes can be effectively prevented from being artificially expanded or lost. The method has important significances for researching population composition features of the plant endophytes and enriching the endophyte resources.

Description

technical field [0001] The invention relates to the technical field of separation of microorganisms, in particular to a method for separation of plant endophytes. Background technique [0002] Plant endophytes refer to fungi or bacteria that live in various tissues and organs of healthy plants. During their life, the host plant does not show external symptoms. Plants and their endophytes are in a mutualistic symbiotic relationship. Plants provide photosynthetic products and minerals for endophytes, and the metabolites of endophytes can stimulate plant growth and development, and improve the resistance of host plants to biotic and abiotic stresses. Due to the unique advantages of plant endophytes as biocontrol factors: they are distributed in different tissues of plants, protected by plant tissues, have a stable ecological environment, and are easier to play a role in biocontrol compared with epiphytic bacteria, so , the research on plant endophytes is becoming the focus of ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12N1/14C12N1/02
CPCC12N1/02C12N1/14C12N1/20
Inventor 陈泽斌夏体渊靳松徐胜光余磊刘佳妮赵凤郭丽红王定康
Owner KUNMING UNIV
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