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A kind of detection method and detection kit of tetracycline

A detection kit and tetracycline technology, applied in the field of analytical chemistry, can solve the problems of long test time, interference detection judgment, poor sensitivity, etc., and achieve the effects of reducing non-specific interference, convenient and rapid detection, and simplified operation

Active Publication Date: 2019-08-27
GUANGDONG INST OF ECO ENVIRONMENT & SOIL SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, microbiological analysis methods often have false positive results, which interfere with detection and judgment; mass spectrometry usually requires complex pretreatment of samples, involving expensive detection instruments, cumbersome operations, long test time, and high costs; traditional enzyme-linked immunoassays, although Simple and fast, but poor sensitivity, difficult to meet the needs of trace detection

Method used

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  • A kind of detection method and detection kit of tetracycline
  • A kind of detection method and detection kit of tetracycline
  • A kind of detection method and detection kit of tetracycline

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0084] A detection method for tetracycline, carried out according to the following steps:

[0085] (1) First use Tris-HCl buffer (20 mM, pH 7.5, containing 150 mM NaCl and 50 mM KCl) to dissolve nucleic acids A, B, C, D, E, and F respectively. 500 nM A was mixed with 500 nM B and reacted at room temperature for 30 minutes to form an A-B mixture. Add tetracycline detection substance and react at room temperature for 45 minutes to complete the displacement reaction and replace B.

[0086] (2) 500 nM C, D, and E were mixed thoroughly, and reacted at room temperature for 30 minutes to form a C-D-E mixture. Add B to the C-D-E mixture, mix thoroughly, and react at room temperature for 50 minutes to complete the replacement reaction. Then add 500 nM F, mix thoroughly, and react at room temperature for 50 minutes to complete the displacement reaction. A large number of C are replaced.

[0087] (3) Add 0.3 M hemin (hemin) and react at room temperature for 30 minutes. Take 50 L of ...

Embodiment 2

[0089] A tetracycline detection kit, comprising the following components:

[0090] (1) Nucleic acid sequences A, B, C, D, E, F. Their sequence is as follows:

[0091] A: 5'-CGTACGGAATTCGCTAGCCCCCCGGCAGGCCACGGCTTGGGTTGGTCC

[0092] CACTGCGCGTGGATCCGAGCTCCACGTG-3' (SEQ ID NO: 1);

[0093] B: 5'-GATCCACGCGCAGTGG (region 4)-GACCAA (region 5)-3' (SEQ ID NO: 2);

[0094] C: 5'-GGGTA (region 1)-GGGCGGGTTGGG (region 2)-3' (SEQ ID NO: 3);

[0095] D: 5'-CCACATACATCATATT (region 6)-CCCT (region 3)-GATCCACGCGCAGTGG (region 4)-3' (SEQ ID NO: 4);

[0096] E: 5'-TTGGTC (region 5*)-CCACTGCGCGTGGATC (region 4*)-AGGG (region 3*)-CCCAACCCGCCC (region 2*)-3' (SEQ ID NO: 5);

[0097] F: 5'-GGGCGGGTTGGG (region 2)-CCCT (region 3)-GATCCACGCGCAGTGG (region 4)-3' (SEQ ID NO: 6).

[0098](2) Tris-HCl buffer containing 150 mM NaCl and 50 mM KCl.

[0099] (3) Hemin.

[0100] (4) Chromogenic buffer system, containing 26.6 mM citric acid, 51.4 mM disodium hydrogen phosphate, 25 mM KCl, 10 L 0.5% T...

Embodiment 3

[0102] Detection of different concentrations of tetracycline:

[0103] Prepare tetracycline standard solutions with concentrations of 10 pM, 100 pM, 1 nM, 10 nM and 100 nM, respectively, and store them at room temperature.

[0104] Tetracycline solutions of different concentrations were added to the reaction system described in Example 1 respectively, and the experimental results were observed after sufficient reaction, such as figure 2 As shown, 10 pM tetracycline produces a distinct blue color change, indicating a detection limit of 10 pM. As the concentration of tetracycline increases, the color also increases and gradually becomes saturated.

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Abstract

The invention discloses a tetracycline detection method and a detection kit. The tetracycline aptamer is adopted as a molecular recognition element, for the design of non-stem-loop structure nucleic acids, when tetracycline is present in the system, a DNA self-assembly can be started, and the amplification of detection signals is achieved, and a large number of G-rich base nucleic acid sequences are substituted out. The finally formed G tetramer has horseradish peroxidase-like catalytic activity, the TMB-H2O2 detection system can be catalytically oxidized, the colorless substrate is turned into blue in color, the concentration of tetracycline and changes in blue color are positively correlated, and the concentration of tetracycline in the detection system can thus be judged. The tetracycline detection method is high in sensitivity, the detection limit of tetracycline is 10 pM, and the detection is excellent in specificity, common interferences do not have an impact on detection. Detection instruments are not required in the detection procedure, the results are directly visible to bare eyes, and the detection kit has the advantages of being simple in operation, low in cost, and rapid in response, the detection method and detection kit are suitable for being widely promoted at the basic level.

Description

technical field [0001] The invention belongs to the field of analytical chemistry, and in particular relates to a tetracycline detection method and a detection kit. Background technique [0002] Tetracycline (Tetracycline) is a broad-spectrum antibiotic, commonly used to prevent poultry diseases, promote the growth of young poultry and so on. Long-term consumption of foods containing tetracycline will cause great harm to human health, cause bone deformities and growth inhibition, interfere with the normal physiological functions of the human body, affect the growth and development of children and adolescents, reduce the body's resistance to pathogenic bacteria, and cause Liver and kidney damage, causing vestibular reaction, allergic reaction, allergic reaction, drug resistance, etc. [0003] At present, the detection of tetracycline residues mainly uses microbial methods, mass spectrometry and enzyme-linked immunoassay. However, microbiological analysis methods often have ...

Claims

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Application Information

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IPC IPC(8): G01N33/535G01N33/53G01N21/78C12N15/115
Inventor 陈俊华李定强
Owner GUANGDONG INST OF ECO ENVIRONMENT & SOIL SCI
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