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Efficient screening method of low-abundance soil indigenous lactic acid bacteria

A technique for indigenous lactic acid bacteria, screening methods, applied in the directions of microorganism-based methods, biochemical equipment and methods, bacteria, etc.

Pending Publication Date: 2022-08-05
至农科技发展(浙江)有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, if indigenous lactic acid bacteria can be isolated from the soil to be improved, and then applied to the soil after in vitro propagation, their colonization ability and comprehensive inhibitory effect on harmful microorganisms in the cultivated land are unmatched by exogenous lactic acid bacteria.

Method used

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  • Efficient screening method of low-abundance soil indigenous lactic acid bacteria
  • Efficient screening method of low-abundance soil indigenous lactic acid bacteria
  • Efficient screening method of low-abundance soil indigenous lactic acid bacteria

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] A kind of high-efficiency screening method of low-abundance soil indigenous lactic acid bacteria of the present invention comprises the following steps:

[0033] S1. Sample collection.

[0034] In a farm in Nanxun District, Huzhou City, Zhejiang Province, 15 kilograms of soil samples were taken from the same location on the farmland where tomatoes have been planted continuously for 3 years. After preliminary mixing, the soil samples were put into 3 sterile sample bags and brought back to the laboratory. Refrigerate at 4°C for later use. The experimental object of this example is the soil sample in one of the sterile sample bags.

[0035] S2, one enrichment culture.

[0036] The configuration method of MRS liquid medium is as follows: Weigh 52.24 g of MRS medium, dissolve it in 1000 mL of distilled water by heating, and sterilize by autoclaving at 118° C. for 15 minutes. Prepare enough MRS liquid medium for backup.

[0037] The composition of the MRS liquid medium af...

Embodiment 2

[0062] The main difference between Example 2 and Example 1 is that the steps of enrichment culture are different. Example 1 carried out three enrichment cultures, while Example 2 carried out only two enrichment cultures. The specific training plan is as follows:

[0063] S1. Sample collection.

[0064] The experimental sample in this example is the soil sample in another sterile sample bag in Example 1.

[0065] S2, one enrichment culture.

[0066] The method was the same as that of Example 1, and the culture time was 22h.

[0067] S3, secondary enrichment culture.

[0068] Prepare a new Erlenmeyer flask, add 300mL MRS liquid medium, take 1mL from each Erlenmeyer flask that has completed the secondary enrichment culture and add it to the new Erlenmeyer flask, and put the new Erlenmeyer flask into the shaker , 37 ℃, 140rmp under the condition of culturing for 10h and then take out.

[0069] S4, directional screening of calcium dissolving circle.

[0070] Using the gradie...

Embodiment 3

[0074] The main difference between Example 3 and Example 1 is that this Example 1 is enriched and cultured only once. The specific training plan is as follows:

[0075] S1. Sample collection.

[0076] The experimental sample in this example is the soil sample in the third sterile sample bag in Example 1.

[0077] S2, one enrichment culture.

[0078] Prepare 20 Erlenmeyer flasks, each with 300 mL of MRS liquid medium, add 50 g of soil samples to each Erlenmeyer flask, and adjust the pH of the MRS liquid medium to 5.7. The conical flask was placed in a shaker, and incubated at 37°C and 140rmp for 30h, and the culture medium was not turbid. Continue to culture until the culture time reaches 48h.

[0079] S3, directional screening of calcium dissolving circle.

[0080] 100 μL of the culture solution was uniformly spread on MRS agar plates containing 1% calcium carbonate, and incubated at 37° C. for 30 h. The results of directional screening of the calcification circle are as...

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Abstract

The invention belongs to the technical field of soil improvement and soil microorganisms, and particularly relates to an efficient screening method of low-abundance soil indigenous lactic acid bacteria, which comprises the following steps: S1, sample collection; s2, primary enrichment culture: adding a soil sample into a plurality of MRS liquid culture media, adjusting the pH value to 5.7 + / -0.2, and culturing for more than or equal to 8 hours; s3, sampling with a double-layer filter membrane, then carrying out secondary enrichment culture, filtering a bacterial liquid obtained after the primary enrichment culture with the double-layer filter membrane, and then respectively putting the bacterial liquid into a plurality of new MRS liquid culture mediums for continuous culture for more than or equal to 8 hours; s4, third-time enrichment culture, wherein a certain amount of culture solution is taken from each bacterium solution obtained after second-time enrichment culture and added into another new MRS liquid culture medium to continue to be cultured for 6 h or longer; s5, directional screening of calcium dissolving rings; and S6, identifying the strain. The screening method of the low-abundance soil indigenous lactic acid bacteria is established, and technical support can be provided for expanding the source of the soil lactic acid bacteria and constructing an efficient lactic acid bacteria resource library.

Description

technical field [0001] The invention belongs to the technical field of soil improvement and soil microorganisms, in particular to an efficient screening method for low-abundance soil indigenous lactic acid bacteria. Background technique [0002] Lactic acid bacteria is a general term for bacteria that use sugars as substrates, produce organic acids such as lactic acid and phenyllactic acid, and secrete bacteriocin and other plant pathogenic bacteria growth inhibitors. It is safe and edible, and is currently the safest agricultural microorganism strain. . Studies have shown that lactic acid bacteria can promote the growth of crops, inhibit the reproduction of plant pathogens, activate the beneficial flora of plants, improve soil quality and fertility, and alleviate the toxicity of heavy metals and pesticides. At present, lactic acid bacteria have become an important component in soil conditioners. Such as Chinese patent document CN 110846041 A discloses a kind of soybean wa...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12N1/02C12R1/01
CPCC12N1/20C12N1/02
Inventor 吴酬飞陈添裕任若琛黄岸城胡婷宋园园张红芳严飞杨王思勤赵心旷谢小玲廖浩然
Owner 至农科技发展(浙江)有限公司
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