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A method for detecting cyromazine in milk based on nucleic acid aptamer

A nucleic acid aptamer and cyromazine technology, which is applied in the preparation of test samples, measuring devices, material excitation analysis and other directions, can solve the problems of high cost, heavy workload, difficulty in satisfying rapid detection, etc., and achieves low cost , Easy operation, high sensitivity effect

Active Publication Date: 2021-07-20
SHANGHAI INST OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

With the detection of cyromazine veterinary drug residues included in raw milk purchases and dairy product factory inspection items, and due to the limitations of the above-mentioned detection methods and conditions, the huge number of detections will lead to huge costs and heavy workloads, making it difficult to meet the requirements. The demand for on-site rapid detection objectively requires the improvement of existing detection methods and the development of a high-throughput, rapid, and highly sensitive detection method for cyromazine for daily monitoring

Method used

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  • A method for detecting cyromazine in milk based on nucleic acid aptamer
  • A method for detecting cyromazine in milk based on nucleic acid aptamer
  • A method for detecting cyromazine in milk based on nucleic acid aptamer

Examples

Experimental program
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Effect test

Embodiment 1

[0042] like figure 1As shown, the method for the rapid detection of cyromazine in milk based on surface-enhanced Raman spectroscopy based on the nucleic acid aptamer sensor is as follows:

[0043] 1) Preparation of detection reagent: prepare ssDNA nucleic acid aptamer solution with a concentration of 2000nmol / L in distilled water, and the sequence of the nucleic acid aptamer is 5'-CCCGGTTGGTTGGTTGGTTTTCCC-3'; prepare a silver nitrate solution with a concentration of 0.1mmol / L in distilled water; Prepare a sodium borohydride solution with a concentration of 0.1 mmol / L in distilled water.

[0044] 2) Pretreatment of milk samples: In a 10 mL centrifuge tube, add 3 mL of 1 wt % trichloroacetic acid solution and 3 mL of milk samples, and let stand for 10 minutes. Then centrifuge at 10000 r / min for 10 minutes, take the supernatant, and adjust the pH=5 with 2.0 mol / L NaOH.

[0045] 3) Draw the standard curves of different concentrations of cyromazine standard solutions and the corr...

Embodiment 2

[0049] The detection process in Example 1 was further optimized, and the Raman intensity of the 0.5ppm cyromazine solution under the conditions of nucleic acid aptamer concentration 0-200nmol / L, pH value 4-11, and reaction time 0-30min was measured respectively , see the results respectively figure 2 , image 3 , Figure 4 shown. It can be seen that the optimum concentration of the nucleic acid aptamer is above 70nmol / L, the optimum reaction pH is 4.5-6, and the optimum reaction time is 7-15min.

Embodiment 3

[0051] A method for detecting cyromazine in milk based on a nucleic acid aptamer, wherein the content of cyromazine in a milk sample is 0-0.5ppm. The method includes the following steps:

[0052] 1) Preparation of detection reagents comprising nucleic acid aptamers: comprising a nucleic acid aptamer solution with a concentration of 70nmol / L, a silver nitrate solution with a concentration of 0.12mmol / L and a sodium borohydride solution with a concentration of 0.08mmol / L; In the ligand solution, the nucleic acid aptamer sequence is 5'-CCCGGTTGGTTGGTTGGTTTTCCC-3'.

[0053] Pretreatment of the milk sample: adding trichloroacetic acid to the milk sample, centrifuging after standing still, taking the supernatant, and adjusting the pH value of the supernatant to 4.5.

[0054] 2) draw the standard curve of the cyromazine standard solution of different concentrations and the corresponding Raman intensity:

[0055] 2-1) adding cyromazine standard solutions of different concentrations ...

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Abstract

The invention relates to a method for detecting cyromazine in milk based on a nucleic acid aptamer, the method comprising the following steps: 1) preparing a detection reagent comprising a nucleic acid aptamer, and pretreating a milk sample; 2) drawing different concentrations The standard curve of the cyromazine standard solution and the corresponding Raman intensity; 3) the detection reagent is added to the pretreated milk sample, and the Raman intensity is measured after the reaction, and the standard curve is used to obtain the cyromazine in the milk sample content data. Compared with the prior art, the present invention uses cyromazine-specific nucleic acid aptamers to control synthesis of nano-silver with Raman effect to detect cyromazine. The detection method has high sensitivity and good selectivity, and the minimum detection limit is 15.5ppb, easy to operate, simple pre-treatment, no need for large instruments, low cost, and can be used for the detection of cyromazine in milk.

Description

technical field [0001] The invention belongs to the technical field of detection of cyromazine in milk, and relates to a method for using a cyromazine nucleic acid aptamer to control and synthesize nano-silver with a Raman effect, and to detect cyromazine in milk by a Raman spectrometer. Background technique [0002] The chemical name of Cyromazine (Cyromazine, Cyr) is 2-cyclopropylamino-4,6-diaminotriazine, which belongs to triazine or The s-triazine compound is a high-efficiency insect growth inhibitor and parasite-killing insecticide, and is used as a feed additive in the process of livestock and poultry breeding. Studies have shown that most of the cyromazine entering the animal body is excreted through milk or feces in the form of the original drug or metabolites, and then exposed to the soil and water environment through livestock and poultry manure. Cyromazine in the soil or water environment re-enters the food chain through environmental fate, causing potential hidd...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/65G01N1/28
CPCG01N1/28G01N21/658
Inventor 邢海波党旭费雪莲龚梓轩孙诗韵胡晓钧
Owner SHANGHAI INST OF TECH
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