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Method of constructing bacillus licheniformis by knocking out ccpN gene, strain and application of strain

A technology of Bacillus licheniformis and Bacillus licheniformis, which is applied in the field of strain transformation of Bacillus licheniformis, and can solve problems affecting the production of bacitracin, not given, and unpredictable

Active Publication Date: 2018-07-13
LIFECOME BIOCHEM
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  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, this prior art does not give the technical inspiration of "the correlation between the transcriptional repressor CcpN and the synthesis of bacitracin", and it is still unpredictable whether the transcriptional repressor CcpN will affect the production of bacitracin

Method used

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  • Method of constructing bacillus licheniformis by knocking out ccpN gene, strain and application of strain
  • Method of constructing bacillus licheniformis by knocking out ccpN gene, strain and application of strain
  • Method of constructing bacillus licheniformis by knocking out ccpN gene, strain and application of strain

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Embodiment Construction

[0024] by knockout ccpN The method for genetically constructing bacillus licheniformis comprises the following steps:

[0025] (1) Using the genomic DNA of Bacillus licheniformis DW2 as a template, PCR amplified ccpN The upstream homology arm of the gene and ccpN The downstream homology arm of the gene; and then use the overlap extension PCR method to ccpN The upstream homology arm of the gene and ccpN The downstream homology arms of the gene are spliced ​​together to obtain the fusion gene sequence A;

[0026] (2) Using restriction endonucleases Xba I and Sac I to perform double enzyme digestion on the fusion gene sequence A obtained in step (1) to obtain the enzyme cut gene sequence A;

[0027] (3) Prepare the plasmid T2(2)-ori, and use restriction endonucleases XbaI and SacI to double-enzyme-digest the plasmid T2(2)-ori to obtain the restriction endonuclease T2(2)-ori;

[0028] (4) Link the enzyme-cut gene sequence A obtained in step (2) to the enzyme-cut plasmid T2(2)...

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Abstract

The invention provides a method of constructing bacillus licheniformis by knocking out a ccpN gene, a strain and application of the strain. By adopting a gene engineeringmethod, the bacillus licheniformis DW2 delta ccpN is constructed by knocking out the ccpN gene in a bacillus licheniformis DW2 genome; and compared with the bacillus licheniformis DW2, for the strain, the yield of bacitracin is increased by 16 percent or above.

Description

technical field [0001] The invention relates to the field of Bacillus licheniformis strain transformation, especially a ccpN The method and strain of gene constructing Bacillus licheniformis and its application. Background technique [0002] Bacitracin is a cyclic peptide antibiotic produced by Bacillus subtilis and Bacillus licheniformis. As a broad-spectrum antibiotic, bacitracin can effectively inhibit Gram-positive bacteria and some Gram-negative bacteria. Moreover, bacitracin is hardly absorbed in the intestinal tract of animals, and is excreted quickly without residue, so it is widely used in feed addition. [0003] Bacitracin is a class of cyclic peptide antibiotics composed of 12 amino acid residues. The constituent amino acids of bacitracin include ornithine (Orn), D-phenylalanine (D-Phe), isoleucine (His ), D-Aspartic Acid (D-Asp), Asparagine (Asn), Lysine (Lys), D-Glutamic Acid (D-Glu), Cysteine ​​(Cys), Leucine (Leu), isoleucine (Ile) and valine (Val) 11 kind...

Claims

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Application Information

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IPC IPC(8): C12N1/21C12N15/03C12P21/00C12R1/10
CPCC07K14/32C12N15/03
Inventor 陈守文蔡冬波朱江占杨杨李俊辉段茂华楼丽君陈晓斌
Owner LIFECOME BIOCHEM
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