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Method for isolated culturing of root nodule endophytes of sea buckthorn plants

A seabuckthorn root nodule, isolation and culture technology, applied in the isolation and culture of plant root nodule endophytes, and in the field of isolation and culture of seabuckthorn plant root nodule endophytes, can solve the problem of large nutritional differences, inability to cultivate microorganisms, and small number and types of endophytes and other problems, to achieve the effect of increasing species and quantity, multi-endophyte resources, and shortening time

Inactive Publication Date: 2018-02-16
NORTHWEST NORMAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the large nutritional difference between the culture medium and the natural environment, it is impossible to simulate the original conditions of the habitat of seabuckthorn endophytes, and a large number of microorganisms cannot be cultivated.
The traditional method used PDA medium, beef extract peptone medium and Gaoshi No. 1 medium respectively when isolating and cultivating the endophytic bacteria of seabuckthorn root nodules. The number and type of bacteria in the whole plant

Method used

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  • Method for isolated culturing of root nodule endophytes of sea buckthorn plants
  • Method for isolated culturing of root nodule endophytes of sea buckthorn plants

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0015] (1) Surface disinfection of seabuckthorn root nodule samples

[0016] Select fresh and tender seabuckthorn root nodules, and carry out surface disinfection on seabuckthorn root nodules: take spare seabuckthorn root nodules, rinse them with running tap water in turn, and wash away impurities such as sediment with a brush, then rinse with sterile water for 3 to 5 times , surface disinfection with 75% ethanol for 8-10 minutes, surface disinfection with 1% NaClO solution for 5-7 minutes, and then rinse with sterile water for 3-5 times (avoid damage to the surface of seabuckthorn root nodule samples during the entire operation).

[0017] (2) Isolation and cultivation of endophytes

[0018] Preparation of seabuckthorn root extract: Wash and chop the seabuckthorn root, mix it with distilled water at a volume ratio of 1:1, boil it, simmer for about 1 hour, filter, take the clear liquid, and sterilize it at 121°C for storage for later use.

[0019] Isolation and cultivation of ...

Embodiment 2

[0027] (1) Surface disinfection of seabuckthorn nodule samples: same as in Example 1.

[0028] (2) Isolation and cultivation of endophytes: Isolation and cultivation of endophytes: Cut the root nodules into 1-2 mm thin slices with a sterile scalpel, insert them into three different agar media, culture them at 28°C, and wait for the nodules to grow. After the flake grows bacteria, it is picked onto the plate with an inoculation needle for further purification.

[0029] The endophytic fungus uses PDA medium, and sea buckthorn root extract is added to the PDA medium (the volume ratio of medium to sea buckthorn root extract is 1:15), and the plate is cultured at 28°C for 5 days.

[0030] Beef extract peptone medium was used for endophytic bacteria, and sea buckthorn root extract was added to the beef extract peptone medium (the volume ratio of medium to sea buckthorn root extract was 1:15), and the plate was cultured at 28°C for 5 days.

[0031] The endophytic actinomycetes use G...

Embodiment 3

[0036] (1) Surface disinfection of seabuckthorn nodule samples: same as in Example 1.

[0037] (2) Isolation and cultivation of endophytes: Isolation and cultivation of endophytes: Cut the root nodules into 1-2mm thin slices with a sterile scalpel, insert them into three different agar mediums, culture them at 30°C, and wait for the nodules to grow. After the flake grows bacteria, it is picked onto the plate with an inoculation needle for further purification.

[0038] The endophytic fungus uses PDA medium, and sea buckthorn root extract is added to the PDA medium (the volume ratio of medium to sea buckthorn root extract is 1:18), and the plate is cultured at 30°C for 3 days.

[0039] Beef extract peptone medium was used for endophytic bacteria, and sea buckthorn root extract was added to the beef extract peptone medium (the volume ratio of medium to sea buckthorn root extract was 1:15), and the plate was cultured at 30°C for 3 days.

[0040] The endophytic actinomycetes use ...

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Abstract

The invention discloses a method for isolated culturing of root nodule endophytes of sea buckthorn plants. The method comprises the steps as follows: a sea buckthorn root nodule sample is subjected tosurface disinfection, and with adoption of a root nodule slicing method and a flat plate isolation method, sea buckthorn root leachate is added to different media for isolated culturing of differentsea buckthorn root nodule endophytes. Research shows that when the sea buckthorn root nodule endophytes are isolated, the media are more nutritious after the sea buckthorn root leachate is added and more suitable for growth of the endophytes. During isolation of the endophytes, the bacteria lawn growing faster overspreads around root nodule slices, bacteria and actinomycetes growing slowly are hindered, especially growth of Frankiaceae; besides, during expanded culture of strains in the later period, the media with the sea buckthorn root leachate can also obviously promote growth of the strains, the variety and the quantity of the root nodule endophytes that can be cultured are increased obviously, more endophyte resources are obtained, and isolated culturing time of the endophytes can beshortened.

Description

technical field [0001] The invention relates to a method for isolating and cultivating endophytic bacteria in plant root nodules, in particular to a method for isolating and cultivating endophytic bacteria in root nodules of plants belonging to the genus Hippophae, belonging to the technical field of microorganism isolation and culture. Background technique [0002] Endophytes are a large group of microorganisms that live in the tissues and organs of healthy plants at certain or all stages, and they can form parasitic, symbiotic, saprophytic and other relationships with plants. Studies have found that not only endophytes can be isolated from various plants, but also endophytes are found in the roots, stems, leaves, flowers, fruits and seeds of the same plant. The number of endophytic bacteria is huge and there are many types, including endophytic fungi, endophytic bacteria and endophytic actinomycetes. Plant endophytes are rich in biodiversity. For a plant, there are usuall...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12N1/02C12R1/41
CPCC12N1/02C12N1/20
Inventor 张爱梅韩雪英
Owner NORTHWEST NORMAL UNIVERSITY
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