Nocardia seriolae attenuated strain and preparation method and application thereof

A technology of Nocardia amberjack and an attenuated strain is applied in the field of genetic engineering, and can solve the problems of antibiotic resistance, accumulated poisoning in the human body, and long treatment course of treatment, and achieves a short cycle, reduced pathogenicity, and improved preparation. simple method effect

Active Publication Date: 2021-05-04
SHENZHEN INST OF GUANGDONG OCEAN UNIV +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] In recent years, fish nocardiosis has attracted the attention of more and more scholars. It is difficult to cure bacterial infectious diseases with antibiotics. Du...

Method used

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  • Nocardia seriolae attenuated strain and preparation method and application thereof
  • Nocardia seriolae attenuated strain and preparation method and application thereof
  • Nocardia seriolae attenuated strain and preparation method and application thereof

Examples

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Embodiment 1

[0027] This embodiment provides an attenuated strain of Nocardia japonica, which is a Nocardia japonica strain containing a deletion of the alanine dehydrogenase gene; the nucleotide of the alanine dehydrogenase gene The sequence is shown in the sequence listing SEQ ID NO: 1, the amino acid sequence of the alanine dehydrogenase is shown in the sequence listing SEQ ID NO: 12, wherein the nucleotides of the upstream fragment of the alanine dehydrogenase gene The sequence is shown in SEQ ID NO:2 in the sequence listing, and the nucleotide sequence of the downstream fragment of the alanine dehydrogenase gene is shown in SEQ ID NO:3 in the sequence listing.

[0028] Specifically, the preparation method of the attenuated Nocardia strain of yellowtail comprises the following steps:

[0029] S1. Construction of a homologous recombination vector: aseptically inoculate the preserved Nocardia spp. ZJ0503 strain into Brain Heart Infusion (BHI) solid medium (purchased from Guangdong Huanka...

Embodiment 2

[0047] This embodiment provides an attenuated strain of Nocardia japonica, which is a Nocardia japonica strain containing a deletion of the alanine dehydrogenase gene; the nucleotide of the alanine dehydrogenase gene The sequence is shown in the sequence listing SEQ ID NO: 1, wherein the nucleotide sequence of the upstream fragment of the alanine dehydrogenase gene is shown in the sequence listing SEQ ID NO: 2, and the downstream fragment of the alanine dehydrogenase gene The nucleotide sequence is shown in the sequence listing SEQ ID NO:3.

[0048] Specifically, the preparation method of the attenuated Nocardia strain of yellowtail comprises the following steps:

[0049] S1. Construction of a homologous recombination vector: aseptically inoculate the preserved Nocardia spp. ZJ0503 strain into Brain Heart Infusion (BHI) solid medium (purchased from Guangdong Huankai Microbial Technology Co., Ltd. ), cultured upside down at 26°C. After a single colony grows on the plate, pick...

Embodiment 3

[0055] This embodiment provides an attenuated strain of Nocardia japonica, which is a Nocardia japonica strain containing a deletion of the alanine dehydrogenase gene; the nucleotide of the alanine dehydrogenase gene The sequence is shown in the sequence listing SEQ ID NO: 1, wherein the nucleotide sequence of the upstream fragment of the alanine dehydrogenase gene is shown in the sequence listing SEQ ID NO: 2, and the downstream fragment of the alanine dehydrogenase gene The nucleotide sequence is shown in the sequence listing SEQ ID NO:3.

[0056] Specifically, the preparation method of the attenuated Nocardia strain of yellowtail comprises the following steps:

[0057] S1. Construction of a homologous recombination vector: aseptically inoculate the preserved Nocardia spp. ZJ0503 strain into Brain Heart Infusion (BHI) solid medium (purchased from Guangdong Huankai Microbial Technology Co., Ltd. ), cultured upside down at 30°C. When a single colony grows on the plate, pick ...

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Abstract

The invention is applicable to the technical field of genetic engineering and provides a nocardia seriolae attenuated strain and a preparation method and application thereof. The nocardia seriolae attenuated strain is a nocardia seriolae strain containing alanine dehydrogenase gene deletion. The preparation method comprises the following steps of: connecting an upstream fragment and a downstream fragment of an alanine dehydrogenase gene into a vector to obtain a recombinant vector; taking a nocardia seriolae wild strain to prepare nocardia seriolae competent cells; performing electrotransformation on the nocardia seriolae competent cells by utilizing the recombinant vector to obtain electrotransformation bacterial liquid; and culturing and screening the electrotransformation bacterial liquid to obtain the nocardia seriolae attenuated strain. The nocardia seriolae attenuated strain provided by the invention can be used for preventing nocardiosis infection; after the alanine dehydrogenase gene is deleted by a gene knockout method, the pathogenicity of bacteria can be effectively reduced, but relatively good immunogenicity is still retained.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, and in particular relates to an attenuated strain of Nocardia japonica and its preparation method and application. Background technique [0002] Yellowtail Nocardia is the main pathogen of fish nocardiosis, which belongs to Bacteria, Firmicutes, Actinobacteria, Actinomycetes ( Actinobacterales), Nocardiaceae, Nocardia. Fish nocardiosis is extremely harmful to the aquaculture industry. Nocardiosis occurs frequently in farmed fish in Asia. The affected species include amberjack, pomfret, large yellow croaker, lucky fish, tilapia, and snakehead , largemouth bass, grouper, four-finger threadfin, purple snapper, red snapper, six-banded trevally, etc. With the improvement of the intensification of the aquaculture industry, the diseases in the aquaculture process are also becoming more and more serious. The natural incidence of fish nocardiosis can reach 15% to 30%, and the serious one can...

Claims

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Application Information

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IPC IPC(8): C12N1/21C12N15/74A61K39/05A61P31/04C12R1/365
CPCC12N9/0016C12N15/74C12Y104/01001A61K39/05A61P31/04A61K2039/522A61K2039/552
Inventor 夏立群侯素莹鲁义善王文基陈国权
Owner SHENZHEN INST OF GUANGDONG OCEAN UNIV
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