Construction and usage of lactobacillus single-plasmid Nisin inducible expression vector

Abstract

The invention provides construction and usage of lactobacillus single-plasmid Nisin inducible expression vector, which belongs to the field of biotechnology. The construction comprises: a, acquisition and identification, the preparation of chromosome DNA producing Nisin lactococcus lactis, the design and synthesis of primer, the PCR amplification, recovery, pGEM-T connection and transformation ofa nisRK gene, and the identification of the nisRK gene; and b, the construction of pW425N, the recovery, connection and identification of the nisRK gene and a basic vector pW425et, the acquisition and identification of inducible promoter nisA gene, and the construction of pW425N. The usage comprises: a, the acquisition and identification of a green fluorescent protein gfp gene; b, the construction and identification of a recombinant expression vector; c, the determination of the inducible expression function of a Nisin inducible expression vector for green fluorescent protein; and d, the detection of lactobacillus stability. The invention has the advantages of avoiding the troublesome operation of integrating chromosomes of host bacteria in the prior system and simplifying the construction of recombinant genetic-engineering lactic acid bacteria.

Description

Technical field: [0001] The invention belongs to the field of biotechnology. Background technique [0002] 1 Definition and distribution of lactic acid bacteria [0003] Lactic acid bacteria (LAB) is a general term for a group of bacteria that can ferment carbohydrates to produce large amounts of lactic acid. This type of bacteria is Gram-positive, spherical or rod-shaped, catalase-negative, consumes more than 50% of glucose to produce lactic acid; does not form spores, does not move or has weak motility; can decompose proteins, but not Produce spoilage products; weak ability to decompose fat. [0004] Lactic acid bacteria are widely distributed in nature, including the digestive system, respiratory system, genitourinary system, oral system, skin system and excrement of humans and animals, milk and dairy products, fruits, roots and branches of plants, rotten plants, Fermented animal foods and beverages, compost, soil and sludge, sewage, several clinical samples, etc. [2]...

AI Technical Summary

Problems solved by technology

However, the main drawback is that the host chromosome integration efficiency of the nisRK gene is low, and the same treatment needs to be carried out for each host strain selected. The genetic manipulation is cumbersome, and the vector host spectrum is narrow, which restricts its application.

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