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Isolation and culture method and application of Agrobaterium isolate

A technology for aquaculture and separation culture, applied in the direction of microorganism-based methods, biochemical equipment and methods, medical preparations containing active ingredients, etc., can solve the problem of inhibiting pathogenic bacteria of difficult-to-sea crustaceans, and no seawater crustaceans larvae The launch of probiotic products and other issues

Inactive Publication Date: 2010-04-07
HUAIHAI INST OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] However, there is no relevant seawater crustacean larvae probiotic product available in China at present.
Microbial products such as photosynthetic bacteria and bacillus are mainly used to regulate the water quality of aquaculture water, and it is difficult to have a specific inhibitory effect on seawater crustacean pathogenic bacteria

Method used

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  • Isolation and culture method and application of Agrobaterium isolate
  • Isolation and culture method and application of Agrobaterium isolate
  • Isolation and culture method and application of Agrobaterium isolate

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0066] Example 1. A method for isolating and cultivating Agrobacterium isolate N6 (Agrobacterium isolate N6), the steps of which are: collecting water for breeding mitten crab seedlings, diluting the water gradiently, and coating it on 2216E marine bacteria solid medium, Cultivate at a constant temperature of 20°C for 72 to 96 hours; select strains with different colony forms and inoculate them into 2216E marine bacteria slant medium for storage; then inoculate the slant strains into 2216E liquid medium, 20°C, 160r / min, culture for 48h, and centrifuge Obtain the bacterial thallus; get the bacterial thalline obtained after each centrifugation, and infect the first stage larvae of Eriocheir sinensis mittenus sinensis just out of the membrane through the method of single bacterial cell infection, compare the metamorphosis rate of the second stage larvae of the flealike larvae, resulting in The bacterium with the highest metamorphosis rate of juveniles was the aquaculture probioti...

Embodiment 2

[0067] Example 2. In the isolation and cultivation method described in Example 1, the strain obtained by centrifugation has the following strain characteristics: the strain is a Gram-positive Brevibacterium with a size of 0.2-0.6 μm×0.8-2.0 μm, no spores, and no capsule; On the 2216E marine bacteria plate medium, the shape is milky white, the edge is smooth, and the middle is protruding; the bacterium is an aerobic bacterium, the growth temperature ranges from 0°C to 28°C, the optimum growth temperature is 22°C, and the growth pH range is 5.5 to 11.0 , the optimal pH is 7.6; the NaCl-resistant concentration is 12%, the most suitable salt concentration is 2%, and no NaCl does not grow; the bacteria can utilize glucose but not starch; it can liquefy gelatin; hydrogen sulfide test, V-P test and Hydrogen peroxide contact enzyme test was positive; methyl red test was negative.

Embodiment 3

[0068] Example 3. refer to Figure 1-10 . In the separation and cultivation method described in Example 1 or 2, the fermentation medium of the bacterial strain obtained by centrifugation is: glucose 7g / L, ammonium sulfate 4g / L, yeast extract 1g / L, NaCl 2g / L, pH7.6; The fermentation conditions are: 5% inoculum size, 20° C., 180 r / min, and culture for 24 hours.

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PUM

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Abstract

The invention relates to an isolation and culture method of Agrobaterium isolate N6, which is characterized by comprising the following steps: collecting water which is used for culturing Eriocheir sinensis seedlings, diluting the water gradient and then coating the diluted water gradient on a 2216E solid culture medium, and performing constant-temperature culture for 72-96h at the temperature of 20 DEG C; selecting bacterial strains in different colonial morphologies and inoculating the bacterial strains on a 2261E slope culture medium to be preserved for later use; inoculating the bacterial strains on the slope into a 2216E liquid culture medium for culturing for 48h at the speed of 160r / m under the temperature of 20 DEG C; obtaining bacterial thalli after centrifuging; taking the bacterial thalli obtained after centrifuging; comparing the distortion ratio of zoea in the second stage by a method of infecting zoea of Eriocheir sinensis in the first stage that is just out of film by a single bacterial thallus; and obtaining the Agrobaterium isolate N6, namely the bacterium leading the zoea in the second stage to have the highest distortion ratio. The method for screening probiotic bacteria is simple, clean and efficient, and the probiotic bacteria have health-promoting effects on various shellfish larva, thus enhancing seedling yield greatly.

Description

technical field [0001] The invention relates to a method for isolating and cultivating bacterial strains, in particular to a method for isolating and cultivating Agrobacterium isolate N6 (Agrobacterium isolate N6), an aquaculture probiotic; the invention also relates to the use of the strain. Background technique [0002] In recent years, due to the aggravation of self-pollution of aquaculture environment and the seriousness of natural environment pollution, the living environment of farmed animals has continued to deteriorate, pathogenic microorganisms have increased, and the speed of transmission has accelerated, causing major losses to the aquaculture industry. At the same time, the residues of certain prohibited chemicals used to prevent and treat diseases in animals have greatly affected the quality of products. However, due to the residues of prohibited drugs such as chloromycin and malachite green used to prevent and control diseases in the production process of seawa...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20A61K35/74A61P43/00C12R1/01A61K35/741
Inventor 朱明阎斌伦陈静夏振强
Owner HUAIHAI INST OF TECH
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