Bacillus velezensis DH82 and preparation method and application of antibacterial protein thereof

A technology of Bacillus and DH82, applied in the field of preparation of Bacillus velesi DH82 and its antibacterial protein, can solve the problems of no Bacillus velesi and few prevention and control patents, etc., to improve immunity and protect the body. The effect of healthy growth and good application prospects

Inactive Publication Date: 2019-05-17
THIRD INST OF OCEANOGRAPHY STATE OCEANIC ADMINISTATION
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] At present, there are very few patents on the application of Bacillus velaisi to the prevention and con

Method used

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  • Bacillus velezensis DH82 and preparation method and application of antibacterial protein thereof
  • Bacillus velezensis DH82 and preparation method and application of antibacterial protein thereof
  • Bacillus velezensis DH82 and preparation method and application of antibacterial protein thereof

Examples

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Example Embodiment

[0037] (2) Preparation of fermentation supernatant: inoculate the activated bacterial solution in step (1) into modified LB medium at a rate of 1 to 5% (V / V), 25 to 40°C, 150 to 220 rpm, 10 to 35 hours, All fermentation broths were centrifuged at 5000-12000rpm and 4℃ for 10-30min, and the supernatant was filtered through 0.22μm microporous membrane to obtain the fermentation supernatant;

[0038] (3) Preparation of antibacterial protein: in step (2) 50~2000mL of fermentation supernatant, add solid ammonium sulfate to make the solution saturation of ammonium sulfate reach 40~100%, overnight at 4℃, centrifuge at 6000~10000rpm for 20~30min. The precipitate is dissolved in ultrapure water, dialyzed in a 3000Da dialysis bag for 24 to 48h, and freeze-dried to obtain antibacterial protein.

[0039] The LB medium components used in the examples of the present invention are: 1% peptone, 0.5% yeast powder, 1% sodium chloride, pH 7.3; the modified LB medium components are: 1% tryptone, 0.5% y...

Example Embodiment

[0041] Example 1: Preparation of antibacterial protein

[0042] (1) Fermentation supernatant: Take Velez Bacillus DH82 and inoculate it on modified LB medium at 25°C, 150rpm, 16h, and then inoculate the modified LB medium at 5% (V / V) inoculum. After 35 hours at 25°C, 220 rpm, all fermentation broths were centrifuged at 5000 rpm, 4°C for 30 minutes, and the supernatant was filtered through a 0.22 μm microporous membrane to obtain the fermentation supernatant;

[0043] (2) Antibacterial protein: Add solid ammonium sulfate to the fermentation supernatant to make the ammonium sulfate saturation of the solution reach 40%, overnight at 4°C, centrifuge at 6000rpm for 30min to obtain the precipitate, dissolve it with ultrapure water, dialyze for 24h in a 3000Da dialysis bag, and freeze-dry Get antibacterial protein.

Example Embodiment

[0044] Example 2: Preparation of antibacterial protein

[0045] (1) Fermentation supernatant: Take Velez Bacillus DH82 and inoculate it in modified LB medium, 40℃, 220rpm, 8h, inoculate the modified LB medium at 1% (V / V) inoculum, 40 ℃, 150rpm, 10h, centrifuge all fermentation broth at 12000rpm, 4℃, 10min, take the supernatant and filter through 0.22μm microporous membrane to obtain the fermentation supernatant;

[0046] (2) Antibacterial protein: Add solid ammonium sulfate to the fermentation supernatant to make the ammonium sulfate saturation of the solution reach 100%, overnight at 4°C, centrifuge at 10000rpm for 20min to obtain the precipitate, dissolve it with ultrapure water, dialyze for 48h in a 3000Da dialysis bag, and freeze dry Get antibacterial protein.

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Abstract

The invention discloses a bacillus velezensis DH82 which is deposited on December 4, 2018 in China General Microbiological Culture Collection Center with the preservation number of CGMCC No. 16865. The method further discloses a preparation method of an antibacterial protein thereof by using the bacillus velezensis DH82 and the application of the antibacterial protein in an aquaculture feed additive. The antibacterial protein can effectively prevent and control diseases of aquaculture animals caused by aquatic pathogenic bacteria, thereby protecting the healthy growth of aquatic animals. The protein can be used as a feed additive for fish farming to significantly improve the immunity of the aquatic animals and has a good application prospect.

Description

technical field [0001] The invention relates to a microbial strain and application thereof, in particular to a preparation method and application of a Bacilus velezensis DH82 and an antibacterial protein thereof. Background technique [0002] In recent years, with the rapid development of marine fishery and aquaculture, the purification ability of aquaculture water in my country has weakened, the water environment has been polluted, and pathogenic bacteria have proliferated, which has greatly increased the chance of cross-infection of pathogens among aquatic animals. Edwardsiela tarda, Aeromonas hydrophila, Vibrio campbelii and other aquatic bacterial pathogens often infect fish in the breeding process of sea and freshwater fish skin and gills, causing sepsis, posing a greater threat to the aquaculture industry and causing economic losses. Therefore, it is imminent to seek safe, efficient and ecological control methods. [0003] For the prevention and control of bacterial ...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12P21/00A23K20/147A23K50/80C12R1/07
CPCY02A40/70
Inventor 徐长安王青华黄仕新唐旭罗曼
Owner THIRD INST OF OCEANOGRAPHY STATE OCEANIC ADMINISTATION
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