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Non-antibiotics resistant shuttle plasmid expression carrier and its constructing method and uses

An antibiotic resistance, shuttle plasmid technology, applied in the application, the use of vectors to introduce foreign genetic material, pharmaceutical formulations, etc., can solve the problems of natural ecological environment pollution, and achieve the effect of highlighting environmental protection awareness and effect

Inactive Publication Date: 2004-03-17
王春凤 +1
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Problems solved by technology

[0006] In order to overcome the large-area diffusion and popularization of drug-resistant genes caused by the transfer of drug-resistant genes between bacteria through plasmids, and cause serious pollution to the microenvironment of the human body, the environment in animals, and the natural ecological environment, the present invention constructs non-antibiotic-resistant "Food-grade" and "feed-grade" Lactobacillus expression systems for marking and selection pressure, the purpose is to provide a non-antibiotic resistance shuttle plasmid expression vector and its construction method and application

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  • Non-antibiotics resistant shuttle plasmid expression carrier and its constructing method and uses
  • Non-antibiotics resistant shuttle plasmid expression carrier and its constructing method and uses
  • Non-antibiotics resistant shuttle plasmid expression carrier and its constructing method and uses

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Embodiment Construction

[0035] According to the type and position of the restriction endonucleases on both sides of the antibiotic resistance gene, primers are designed to amplify the non-resistance selection marker candidate gene. First, the basic plasmid vector pMG36e is digested with endonuclease EcoR I at position 2476 to make The circular plasmid vector becomes linear, and the linear pMG36e plasmid vector and ThyA gene are treated with polymerase T4 DNA, and A / T tails are added to the cut, and then the ThyA gene is connected to the linear vector containing the antibiotic resistance gene Circularization, and finally the antibiotic resistance gene was completely removed with restriction enzymes Nsi I and EcoR I.

[0036] (1) Digest the plasmid vector pMG36e with the endonuclease EcoR I at position 2476 to make it linear, and then use T4 DNA polymerase to treat the plasmid vector digested with EcoR I to be linear, so as to facilitate the next step with the foreign fragment to connect.

[0037] (2)...

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Abstract

The present invention belongs to the field of bioengineering technology and is non-antibiotic resistant shuttle plasmid expression vector and its constitution method and application. Basic plasmid carrier pMG36e is enzyme incised in its incising site with endoenzyme EcoRI to become linear. Non-antibiotic resistant gene is connected to the near erythromycin Er end before cyclization into cycle; and the erythromycin gene Er is then incised with endoenzyme on the cycle to form non-antibiotic resistant shuttle plasmid expression vector. The protective antigen gene on human or animal disease inconnected to the non-antibiotic resistant shuttle plasmid expression vector, so as to produce functional food, bacterial suspension with specific disease preventing and treating effect, oral tonic, feed product and animal vaccine. The gene of the present invention may be used in replacing erythromycin gene without problem of antibiotic resistant gene diffusion.

Description

technical field [0001] The invention belongs to the technical field of bioengineering, and relates to a non-antibiotic resistance shuttle plasmid expression vector and a non-antibiotic gene selection pressure transformation method for the antibiotic gene as a selection pressure plasmid carrier. The method is used to construct a non-antibiotic gene selection pressure plasmid expression vector. And the carrier is applied to microorganisms of Lactobacillus family, especially microorganisms of the genus Lactobacillus. Through the carrier and its application in microorganisms belonging to the genus Lactobacillus, it can be used to prevent and treat specific diseases or exert its growth regulation function and nutritional function. Specifically, the present invention relates to the non-antibiotic gene selection pressure transformation scheme and the plasmid vector constructed with this scheme and the ingestible support prepared by using the non-antibiotic gene selection pressure pla...

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Application Information

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IPC IPC(8): A23C9/123A23K10/18A61K35/744A61P1/00C12N1/21C12N15/64C12N15/65C12N15/70C12N15/74
Inventor 王春凤秦泽荣孙哲刘尚高
Owner 王春凤
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