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Engineering lambda bacteriophage capable of specifically killing enterohemorrhagic Escherichia coli and construction method and application of engineering lambda bacteriophage

An enterohemorrhagic, Escherichia coli technology, applied in the direction of bacteriophage, virus/phage, microorganism-based methods, etc., to achieve high specific killing effect and overcome the effect of broad-spectrum bactericidal

Pending Publication Date: 2022-06-10
HENAN NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the widespread use of antibiotics, on the one hand, has led to an increase in the frequency of drug-resistant pathogenic bacteria, on the other hand, the broad-spectrum antibacterial effect of antibiotics will have a lethal effect on all bacteria, and this non-selective killing will kill intestinal bacteria. In situ bacteria, causing the original intestinal ecological environment disorder, this disordered intestinal environment is more conducive to the invasion of pathogenic bacteria, resulting in more adverse conditions
In recent years, studies on intestinal flora have shown that there is a great relationship between intestinal flora and the health of the host. This intestinal micro-ecological environment disrupted by antibiotics will also bring other potential risks to the health of the host. risk

Method used

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  • Engineering lambda bacteriophage capable of specifically killing enterohemorrhagic Escherichia coli and construction method and application of engineering lambda bacteriophage
  • Engineering lambda bacteriophage capable of specifically killing enterohemorrhagic Escherichia coli and construction method and application of engineering lambda bacteriophage
  • Engineering lambda bacteriophage capable of specifically killing enterohemorrhagic Escherichia coli and construction method and application of engineering lambda bacteriophage

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Embodiment

[0039] 1. Genetic engineering of lambda bacteriophage

[0040] The lyase gene of the lambda phage is knocked out and the CRIS PR-Cas3 system-related genes Cas3, CasA, CasB, CasC, CasD, CasE, Cas1, Cas2 and synthetic CrRNA are inserted into the lambda phage to obtain an engineered lambda phage, The CRISPR-Cas3 system was integrated into the host enterohemorrhagic Escherichia coli through lysogeny of λ phage, and the complementary base sequence was mediated by CrRNA to perform specific bactericidal function. The nucleotide sequence of the CRISPR-Cas3 system related gene is as follows: Shown in SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7 and SEQ ID NO:8, The nucleotide sequence of the CrRNA gene is shown in SEQ ID NO:9.

[0041] The specific construction process of the engineered lambda phage is:

[0042] Step S1: Inducing the release of lysogenic lambda phage

[0043] The lysogenic bacteria of λ phage are enriched and cultivated, ...

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Abstract

The invention discloses an engineering lambda bacteriophage capable of specifically killing enterohemorrhagic Escherichia coli and a construction method and application thereof, the engineering lambda bacteriophage is capable of specifically removing enterohemorrhagic Escherichia coli in a complex microbial environment, and a lyase gene of the engineering lambda bacteriophage is knocked out and is loaded with a CRISPR-Cas3 system. The CRISPR-Cas3 system plays a role in killing enterohemorrhagic Escherichia coli with high specificity by utilizing the soluble source characteristic of the engineering lambda bacteriophage, so that the defect of broad-spectrum sterilization of antibiotic and wild bacteriophage treatment is effectively overcome.

Description

technical field [0001] The invention belongs to the technical fields of microbiology, genome engineering and molecular biology, and relates to the application of genetic engineering transformation of lysogenic phages to enhance specific bactericidal ability, in particular to an engineered lambda phage capable of specifically killing enterohaemorrhagic Escherichia coli and its construction method and application. Background technique [0002] Enterohemorrhagic Escherichia coli is a subtype of Escherichia coli, Gram-negative short bacilli with blunt ends, mostly existing alone or in pairs, but not arranged in long chains; the bacteria have a small amount of perinatal flagella, Possesses less athletic ability. Enterohaemorrhagic Escherichia coli is divided into serotypes 157, 26, and 111, and the main pathogenic strain is O157:H7. Its virulence depends on the production of Shiga-like toxins, hemolysins, and adhesion / elimination mainly to the intestinal epithelial barrier (atta...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/01C12N15/70A61K35/76A61P31/04A61P7/04C12R1/92
CPCC12N7/00C12N9/88C12N15/70A61K35/76A61P31/04A61P7/04C12N2795/00021
Inventor 金梦露陈卫华王海磊
Owner HENAN NORMAL UNIV
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