Test strip and method for detecting extractable tetracycline antibiotics in soil by biosensor

A biosensor and tetracycline technology, applied in the field of test strips for extractable tetracycline antibiotics, can solve problems such as tetracycline antibiotics that have not yet been discovered, and achieve the effects of easy promotion and use, simple detection methods, and low cost

Active Publication Date: 2020-08-18
NANJING AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, no method has been found to use whole-cell biosensors to prepare test strips to assess tetracycline antibiotics in soil

Method used

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  • Test strip and method for detecting extractable tetracycline antibiotics in soil by biosensor
  • Test strip and method for detecting extractable tetracycline antibiotics in soil by biosensor
  • Test strip and method for detecting extractable tetracycline antibiotics in soil by biosensor

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Experimental program
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Embodiment Construction

[0058] 1. Preparation of Escherichia coli BL21 / pMTLacZ strain:

[0059] (1) The gene sequence of the pMT fragment (5256bp) of the tetracycline-mediated regulatory system cloned from the plasmid pMTmCherry into Staphylococcusrostri RST11: Tn916 and the transposon Tn10 by using primer polymerase chain reaction (PCR), the annealing temperature is 55 ℃ ( figure 1 ). The plasmids used are shown in Table 1.

[0060] Bacteria and plasmids used in Table 1

[0061]

[0062] The forward primer sequence and reverse primer sequence of the pMT primer are shown in SEQ ID No.1 and SEQ ID No.2,

[0063] SEQ ID No. 1: TCGTGCCAGCTGCATTAAT

[0064] SEQ ID No. 2: TTCACTTTTCTCTATCACTGATAGG;

[0065] (2) Cloning the gene sequence of the LacZ (368bp) reporter gene fragment from the pCU19 plasmid by using primer polymerase chain reaction (PCR), the annealing temperature is 65°C; the gene sequence of the T7 fragment (220bp) cloned in the plasmid pMTmCherry, annealed The temperature is 60°C; th...

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Abstract

The invention discloses a test strip and a method for detecting extractable tetracycline antibiotics in soil by using a biosensor. The test strips include filter paper strips, fixatives, sensitizers and tetracycline biosensor bacterial agents, the test paper is cut from commercially purchased common filter paper, and the tetracycline biosensor bacterial agents are Escherichia coli BL21 bacterial strains carrying pMTLacZ Plasmid (ie Escherichia coli BL21 / pMTLacZ strain). The pMTLacZ plasmid is a synthetic plasmid independently designed by the inventor and can be induced by a variety of tetracycline antibiotics. The pMTLacZ plasmid can produce different amounts of β-galactosidase under the induction of different concentrations of tetracycline, and can degrade the β-galactosidase fixed on the test strip. X‑gal chromogenic reagent, visually displays the concentration of tetracycline antibiotics in the sample. The method for simultaneously detecting six extractable tetracyclines in soil by using the test strip of the invention is simple, fast, intuitive, accurate, easy to carry, environmentally friendly, wide in scope of application, low in cost and easy to popularize and use.

Description

technical field [0001] The invention belongs to the technical field of biological analysis and detection, and in particular relates to a test strip and a method for detecting extractable tetracycline antibiotics in soil by using a biosensor. Background technique [0002] With the increasing degree of industrialization and the sudden occurrence of various chemical products polluting soil, the number of various chemical substances has increased sharply, which has a great impact on the living environment of human beings. It is urgent to identify the acute toxicity of the increasing environmental pollutants. [0003] Tetracyclines have broad-spectrum and high-efficiency bactericidal and antibacterial effects, can be mixed with most drugs or feed additives, and have been widely used in livestock and poultry production. However, excessive use inevitably causes serious drug residues in livestock and poultry manure, which enters the soil environment through manure fertilization, bec...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/18C12N15/70C12R1/19
CPCC12N15/70C12Q1/18G01N2333/245
Inventor 高彦征马钊
Owner NANJING AGRICULTURAL UNIVERSITY
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