Bacteriophage depolymerase capable of degrading Klebsiella pneumoniae capsular polysaccharides and biofilms

A Klebsiella, phage technology, applied in the direction of virus/phage, hydrolase, microorganism, etc., can solve problems such as difficult to remove

Active Publication Date: 2019-03-29
JILIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In addition, biofilm can be stably attached to the surface of medical devices, and it is difficult to remove by conventi...

Method used

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  • Bacteriophage depolymerase capable of degrading Klebsiella pneumoniae capsular polysaccharides and biofilms
  • Bacteriophage depolymerase capable of degrading Klebsiella pneumoniae capsular polysaccharides and biofilms
  • Bacteriophage depolymerase capable of degrading Klebsiella pneumoniae capsular polysaccharides and biofilms

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Embodiment 1

[0026] 1) The 32nd open reading frame from Klebsiella pneumoniae phage GH-K3 was cloned by means of PCR, double enzyme digestion and ligated molecular cloning. depo32 Gene ligation into pET28a plasmid: Restriction sites Nde I and xho I;

[0027] 2) The pET28a- depo32 The plasmid was transformed into Escherichia coli BL21(DE3), and BL21- depo32 Escherichia coli;

[0028] 3) Insert BL21- depo32 In 1L of LB liquid medium containing 50 μg / ml kanamycin, shake culture at 37°C until the logarithmic growth phase (OD 600 0.6~1.0);

[0029] 4) Add isopropyl-β-D-thiogalactoside to the culture medium to a final concentration of 1 mM, and shake at 16°C for 16 hours at 180 rpm;

[0030] 5) will induce the expression of BL21- depo32 After the bacterial liquid is collected by centrifugation, the bacteria are collected and centrifuged, and then ultrasonically crushed;

[0031] 6) Gently add the supernatant sample to the equilibrated Ni-NTA affinity chromatography column, was...

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Abstract

The invention discloses a bacteriophage depolymerase capable of degrading Klebsiella pneumoniae capsular polysaccharides and biofilms. By prokaryotic expression and purification of a 32nd open readingframe in a genome of Klebsiella pneumoniae bacteriophage GH-K3, a brand-new depolymerase depo32 with high activity is obtained. The depolymerase has high-efficiency degradation effect on the capsularpolysaccharides and biofilms formed by Klebsiella pneumoniae.

Description

technical field [0001] The invention discloses a phage depolymerase for degrading Klebsiella pneumoniae capsular polysaccharides and biofilms, relates to a method for degrading biofilms formed by Klebsiella pneumoniae by using phage depolymerization enzymes, and belongs to biotechnology field. Background technique [0002] Klebsiella pneumoniae ( Klebsiella pneumoniae ) is a common zoonotic pathogen. In recent years, multidrug-resistant Klebsiella pneumoniae has become one of the most important pathogens of hospital-acquired infections. The bacterium not only poses a huge threat to human public health, but also affects the healthy development of the animal breeding industry to a certain extent. [0003] Most Klebsiella pneumoniae have the ability to synthesize and secrete capsular polysaccharides. Capsular polysaccharides serve as a natural barrier for bacteria to maintain bacterial virulence, adherence, and block penetration of some antibiotics. As an important virulen...

Claims

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Application Information

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IPC IPC(8): C12N7/00C12N15/70C12N9/16A61K38/46A61P31/04
CPCA01N63/00A61K38/465A61P31/04C12N7/00C12N9/16C12N15/70C12Y301/00
Inventor 韩文瑜顾敬敏蔡若鹏程梦珺
Owner JILIN UNIV
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