Bacillus subtilis

A technology of Bacillus subtilis and strains, applied in the direction of bacteria, enzymes, animal feed, etc., can solve the problems of limited development, difficult direct absorption by animals, etc., and achieve the effects of good stability, mild degradation and fermentation conditions, and high enzyme activity

Inactive Publication Date: 2012-06-27
HUNAN AGRICULTURAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0010] Both keratin and collagen are good animal protein resources, but due to their special chemical structure and relatively stable properties, it

Method used

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  • Bacillus subtilis
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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Take the soil where feathers have been piled up for a long time in Hunan Agricultural University, put it into sterile distilled water, mix it, let it stand, and after clarification, take the supernatant and apply it to solid inorganic salt feather medium, culture at 37°C for 2-3 days, and obtain colonies . Pick and isolate the colony obtained by coating with a toothpick, inoculate it in the inorganic salt feather culture solution, culture it at 37°C for 48 hours, observe the degradation degree of the feather, screen out the effective strains, dip it in the culture solution for multiple streak cultures, and then Pick a single colony and culture it in inorganic salt feather culture solution for repeated screening to obtain a pure strain with better degradation effect. Finally, inoculate a single colony in gelatin medium and culture at 37°C for 12 hours to obtain the strain of the present invention.

Embodiment 2

[0029] Embodiment 2: the thalline staining of bacterial strain of the present invention, electron microscope observation, colony morphological feature observation experiment

[0030] The bacterial strain of the present invention is cultured and grown by streaking in LB liquid medium at 37° C., and the colony is round, with a moist surface, rough and opaque, with irregular, white or yellowish edges. Pick single bacterium colony wherein, carry out Gram staining, after Gram staining, be purple, short rod-shaped explanation bacterial strain of the present invention is Gram-positive bacteria, as figure 1 . Observed in the electron microscope, the bacterium is rod-shaped, the size of a single cell is 0.6-0.7×2-3 microns, and the coloring is uniform. uncapsulated, such as figure 2 .

Embodiment 3

[0031] Embodiment 3: 16SrDNA identification and the construction of phylogenetic tree of bacterial strain of the present invention

[0032] In the present invention, the EDTA method is used to extract the genomic DNA of the strain of the present invention, and the 16S rDNA primer F: agtggccattacggccagagtttgatcmtggctcag (as shown in SEQ ID No: 1); R: agaggccgaggcggcctacgggytaccttgttacgactt (as shown in SEQ ID No: 2) is used to obtain the high-fidelity sequence of the dominant strain , the PCR system is: ddH 2 O: 40.0ul, 10×Buffer (Mg 2+ plus): 5ul, d NTPs: 1ul, F: 1.0ul, R: 1.0ul, Taq enzyme: 1ul, DNA template: 1ul, total volume50ul, pre-denaturation at 95°C for 4min, denaturation at 95°C for 30s, annealing at 55°C for 45s, 72 30 cycles of extension at ℃ for 1.5 min, final extension at 72 °C for 15 min, and 4 °C to ∞. After the PCR was completed, a brighter band at about 1500p was detected by electrophoresis, and the QlAquick Gel Extraction Kit was used for gel extraction an...

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Abstract

The invention relates to bacillus subtilis CH-001, which was collected in China Center for Type Culture Collection (CCTCC) on April 8th, 2011. The collection number is CCTCC NO: M 2011111. The bacillus subtilis has the characteristic of degrading keratin in a broad spectrum manner and has mild degradation and fermentation conditions, high enzyme production activity and high stability. The bacillus subtilis can generate high-activity keratinase, can degrade collagen, has antagonism on pathogenic bacteria such as staphylococcus aureus and the like, has the characteristics of probiotics, and is applicable to engineering bacteria development for fermentation production of animal protein feed additives and keratinase. The bacillus subtilis avoids the conflict between high-activity keratinase generated by most microorganisms and pathogenicity of the microorganisms on human or poultry and the broad spectrum characteristic on the keratin, and the antagonism on the related pathogenic bacteria is researched. The bacillus subtilis is a low-carbon, economic, high-efficiency, safe and green biological resource and has huge potential value for creation of economic and social benefits.

Description

technical field [0001] The invention belongs to the field of development and utilization of microbial germplasm resources, and in particular relates to a Bacillus subtilis CH-001 which can efficiently degrade keratin and collagen and has an antagonistic effect on related pathogenic bacteria. Background technique [0002] Keratin is rich in natural resources, with an annual output of more than hundreds of thousands of tons in my country. It mainly exists in various animal feathers, hair, hooves, etc. Among them, poultry feathers contain 75% to 85% of hard keratin, most of the amino acids in feather keratin are hydrophobic amino acids, and the cystine disulfide bond is difficult to be opened by proteolytic enzymes, so the properties of feather keratin are extremely Stable and not easily hydrolyzed in dilute acid and alkali, it cannot be absorbed only by the digestive enzymes in the animal's own digestive tract. [0003] At present, the treatment methods of feather keratin mai...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12N9/56A23K1/16C12R1/125
Inventor 陈金军胡能渊张铁瀚
Owner HUNAN AGRICULTURAL UNIV
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