Probiotic clone, construction method and application of integrated single-copy functional f4 pilus operon gene

Abstract

The present invention relates to the field of biotechnology, in particular to probiotic clones, construction methods and applications that integrate a single-copy functional F4 pilus operon gene, comprising the following steps: pTargetT- nth / wxya ::P tet Construction of F4 recombinant plasmid; through the construction of a probiotic clone strain integrating a single copy of the functional F4 pili operon gene, a non-resistant single copy F4 pili gene integrated clone is obtained. The probiotic cloning strain is used as a probiotic live vaccine candidate strain and a drug for treating post-weaning diarrhea and edema disease of piglets. Compared with the prior art, the beneficial effects of the present invention are as follows: the original plasmid of the Nissle1917 probiotic is removed, and there is no resistance gene; the recombinant bacterium can effectively improve the adhesion performance to pig intestinal passage epithelial cells; orally immunize mice It can produce immune serum, that is, IgG antibody against F4 pili; the serum of mice after oral immunization can significantly reduce F4 + Adhesion of strains to a porcine intestinal passage cell line.

Description

technical field

[0001] The invention relates to the application field of biotechnology, in particular to the chromosomal integration, stable genetic expression and display of exogenous functional F4 pili protein of bacteria. Background technique

[0002] Escherichia coli Nissle1917 is a non-pathogenic probiotic strain that has no known adverse effects on the host and can give different hosts health benefits. The main active ingredient is mainly used for human intestinal health regulation. In clinical application, the probiotic Nissle1917 strain has been modified as a carrier of genetic engineering, and the probiotic has been used in the development and manufacture of vaccines, tumor treatment, health care products, diagnostic preparations and other products. The EcNc clone is a prototype wild strain of Escherichia coli Nissle1917 genetically modified to remove two cryptic plasmids pMUT1 and pMUT2 in its cell, which has the characteristics of carrying more (large) capacity h...

Examples

Embodiment Construction

[0029] The famous probiotic Escherichia coli Nissle 1917 (EcN) is a non-pathogenic E. coli probiotic, which has been proven to give health benefits to different hosts, and E. coli Nissle1917 is a non-pathogenic strain that can give different hosts Health benefits, no known harmful effects on the host were found, and this strain is used as the main active ingredient of a probiotic product called Mutaflor in Europe, which is mainly used for the regulation of human intestinal health. The famous probiotic Escherichia coli Nissle 1917 (EcNc) was deposited in CGMCC, General Microorganism Center of China Committee for Culture Collection of Microbial Cultures, at the Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, and the date of preservation was July 2018. 2. The deposit number is CGMCC No.16045.

[0030] In this experiment, the probiotic Nissle1917 plasmid-free clone EcNc was used as a vaccine carrier to construct...

technical field

[0001] The invention relates to the application field of biotechnology, in particular to the chromosomal integration, stable genetic expression and display of exogenous functional F4 pili protein of bacteria. Background technique

[0002] Escherichia coli Nissle1917 is a non-pathogenic probiotic strain that has no known adverse effects on the host and can give different hosts health benefits. The main active ingredient is mainly used for human intestinal health regulation. In clinical application, the probiotic Nissle1917 strain has been modified as a carrier of genetic engineering, and the probiotic has been used in the development and manufacture of vaccines, tumor treatment, health care products, diagnostic preparations and other products. The EcNc clone is a prototype wild strain of Escherichia coli Nissle1917 genetically modified to remove two cryptic plasmids pMUT1 and pMUT2 in its cell, which has the characteristics of carrying more (large) capacity h...

Examples

Embodiment Construction

[0029] The famous probiotic Escherichia coli Nissle 1917 (EcN) is a non-pathogenic E. coli probiotic, which has been proven to give health benefits to different hosts, and E. coli Nissle1917 is a non-pathogenic strain that can give different hosts Health benefits, no known harmful effects on the host were found, and this strain is used as the main active ingredient of a probiotic product called Mutaflor in Europe, which is mainly used for the regulation of human intestinal health. The famous probiotic Escherichia coli Nissle 1917 (EcNc) was deposited in CGMCC, General Microorganism Center of China Committee for Culture Collection of Microbial Cultures, at the Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, and the date of preservation was July 2018. 2. The deposit number is CGMCC No.16045.

[0030] In this experiment, the probiotic Nissle1917 plasmid-free clone EcNc was used as a vaccine carrier to construct...

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