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Bacillus thuringiensis capable of producing cellulase and inhibiting vibrio splendidus and using method

A technology of Bacillus aureus and spore thuringiensis, applied in the field of biology, can solve the problems of no reported pathogenic bacteria and secreted cellulase, and achieve the effects of avoiding economic loss, improving utilization rate and reducing loss

Active Publication Date: 2018-11-20
QINGDAO AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is no report that can inhibit the main pathogenic bacteria and the strains that secrete cellulase at the same time.

Method used

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  • Bacillus thuringiensis capable of producing cellulase and inhibiting vibrio splendidus and using method
  • Bacillus thuringiensis capable of producing cellulase and inhibiting vibrio splendidus and using method
  • Bacillus thuringiensis capable of producing cellulase and inhibiting vibrio splendidus and using method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Example 1: Isolation and screening of strains

[0022] 1.1 Isolation of beneficial bacteria

[0023] (1) Initial screening:

[0024] Take 1g of the sample to be screened (soil or sediment), add 99ml of sterile water to prepare a suspension, and then dilute it to 10 -3 , 10 -4 , 10 -5 , 10 -6 , And then spread it evenly on the carboxymethyl cellulose solid medium plate in the ultra-clean workbench, place it in a constant temperature incubator at 30°C and place it upright for 1 hour, and then invert it to grow a single colony.

[0025] Inoculate the cultured strains on another medium. Pour a certain amount of 1mg / ml Congo red solution into the cultured plate to cover the surface of the plate. After dyeing for 1 hour, pour the Congo The red solution is soaked and eluted with 1mol NaCl solution, and a clear transparent circle appears, which is a cellulase-producing strain. A large transparent circle has a strong enzyme production capacity.

[0026] The initial screening medium formu...

Embodiment 2

[0030] Example 2: Identification of strains

[0031] The total DNA of the bacterial liquid was extracted and used as a PCR template of the bacterial liquid, and PCR amplification was carried out with 27F and 1492R primers (Table 2-1). PCR reaction system (10μl): cDNA 1μl, 10×Buffer 1μl, 2.5mM dNTP 0.8μl, 10μM forward and reverse primers 0.5μl each, 0.05μl rTaq enzyme, 2H2O 6.15μl.

[0032] PCR primers:

[0033]

[0034] The PCR reaction conditions are:

[0035]

[0036] After the entire reaction, the reaction product was detected by 1.0% agarose gel electrophoresis. The target band was cut back under the gel imaging system, and DNA was recovered and purified according to the instructions of the Shanghai SanPrep column kit. Recovered DNA is stored at -20°C or used for transformation link. The recovered product was ligated with the PMD18-T cloning vector and ligated overnight at 16°C to obtain the target strain gene sequence PMD18-T recombinant vector.

[0037] The connection system i...

Embodiment 3

[0043] Example 3: Strain activation conditions

[0044] The Bacillus thuringiensis strain is inoculated into an LB medium and cultured at a temperature of 25-30° C. for 8-10 hours to obtain activated Bacillus thuringiensis. The formula of the LB medium is: peptone 10g / L, yeast extract 5g / L, and NaCl 10g / L.

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Abstract

The invention provides a bacillus thuringiensis strain XW008. The strain is collected in the China General Microbiological Culture Collection Center, has a collection number of CGMCC No. 15647, and has a collection date of April 26, 2018. The invention further discloses application of the bacillus thuringiensis. The bacillus thuringiensis is applied to culture of stichopus japonicas, and the application comprises two methods: (1) fermenting stichopus japonicas feed by adopting the bacillus thuringiensis (2) splashing bacterium liquid of the bacillus thuringiensis into a stichopus japonicas culture pond. On one hand, the strain is capable of secreting extracellular cellulase, and the cellulase is capable of decomposing cellulose in the feed; on the other hand, the main pathogen, namely vibrio splendidus, of skin ulcerative syndrome of stichopus japonicas (also called rotten-skin disease) can be effectively inhibited.

Description

Technical field [0001] The invention belongs to the field of biology, and relates to a Bacillus thuringiensis strain, in particular to a Bacillus strain that produces cellulase and inhibits Vibrio and a method of use. Background technique [0002] With the rapid development of the aquaculture industry, the scale of high-density intensive aquaculture is increasing, and the deterioration of the aquaculture environment and the harm of diseases have become more and more obvious to the economic loss of the aquaculture industry. The traditional solution is to use antibiotics; as we all know, the use of antibiotics is prone to the problem of resistant strains and will remain in the body. Probiotics are living microorganisms that can have beneficial effects on animals within a certain concentration range. Probiotics play a role in promoting the growth of animals by enhancing the inhibitory effect of animals on harmful microbial communities in the intestine, or by enhancing non-specific ...

Claims

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Application Information

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IPC IPC(8): C12N1/20A01K61/30A01K61/13A23K50/80A23K10/12C12R1/07
CPCA23K10/12A23K50/80A01K61/13A01K61/30C12N1/20C12N1/205C12R2001/07Y02A40/81
Inventor 冯政夫朱伟赵兰廷胡彦江
Owner QINGDAO AGRI UNIV
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