Method for animal ubiquitous expression of recombinant human lysozyme

A human lysozyme, systemic technology, applied in the field of obtaining new genes, can solve problems such as inability to achieve physical expression

Pending Publication Date: 2017-06-27
FOSHAN UNIVERSITY
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  • Abstract
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  • Claims
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Problems solved by technology

Even so, the human lysozyme obtained by the prior art is mainly expressed in the mammary gland, and cannot be expressed in multiple parts of the body

Method used

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  • Method for animal ubiquitous expression of recombinant human lysozyme
  • Method for animal ubiquitous expression of recombinant human lysozyme
  • Method for animal ubiquitous expression of recombinant human lysozyme

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 3

[0080] Example 3: Detection of biological activity of recombinant human lysozyme in serum

[0081] A. Agar plate method

[0082]Take 100uL Micrococcus lyticus respectively ( M. Lysodeikticus ) evenly spread on the 1.5% agar base broth culture plate, and put filter paper sheets with 10uL serum on the culture medium, the serum of ordinary mice is the negative control (NC), and the positive control is the standard product of human lysozyme. PC is 1ug human lysozyme standard. Set up 4 repetitions, culture at 28°C for 36 hours, observe the diameter of the inhibition zone, and visually judge the expression level and activity of recombinant human lysozyme ( image 3 .C).

[0083] The results showed that the serum samples of No. 1-7 F2 generation transgenic mice could all produce obvious inhibition zones, indicating that the recombinant human lysozyme had biological activity.

[0084] , Nephelometric method

[0085] Micrococcus lyticus was cultured in liquid medium for 24 hours,...

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Abstract

The invention discloses a method for animal ubiquitous expression of recombinant human lysozyme. The method is characterized by comprising the following steps: I, performing human lysozyme BAC cloning, namely, selecting human lysozyme of the Genome Systems company as target hLZ BAC cloning, wherein the GenBank number of the human lysozyme is RP11-72P21; II, removing redundant gene CPSF6, namely, to avoid interference of the redundant gene CPSF6 to the hLZ gene, removing the redundant gene CPSF6 by using a recombinant method, thereby obtaining the human lysozyme pBAC-hLZ-Neo of which the redundant gene is removed. Compared with the prior art, the method has the advantages that the human lysozyme is expressed at multiple parts of bodies. The recombinant human lysozyme expressed in different tissue organs all have bioactivity, the number of bifidobacteria can be remarkably increased (P is less than 0.001) due to the recombinant human lysozyme expressed in intestinal tracts, and meanwhile the number of salmonella can be remarkably reduced (P is less than 0.001).

Description

technical field [0001] The present invention relates to a technique for obtaining new genes through recombination. Background technique [0002] Lysozyme, also known as mureinase or N-acetyl murein hydrolase, is an alkaline enzyme that can hydrolyze mucopolysaccharides in pathogenic bacteria. Mainly by destroying the β-1,4 glycosidic bond between N-acetylmuramic acid and N-acetylglucosamine in the cell wall, the insoluble mucopolysaccharides in the cell wall are decomposed into soluble glycopeptides, resulting in the escape of the contents of the cell wall Bacteria dissolve. Lysozyme can also directly combine with negatively charged viral proteins, and form double salts with DNA, RNA, and apoproteins to inactivate the virus. Therefore, the enzyme has antibacterial, anti-inflammatory, antiviral and other effects. [0003] At present, the lysozyme that is closely related to human life is mainly egg white lysozyme, which is widely derived from poultry egg white. It has a wid...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/85C12N9/36
CPCA01K2227/105A01K2267/01C12N9/2462C12N15/8509C12N2800/107C12Y302/01017
Inventor 刘燊鲁丹王晔张辉华林树茂黄得纯
Owner FOSHAN UNIVERSITY
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