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Nucleic acid aptamer specifically bound with vomitoxin, preparation method and application

A technology of nucleic acid aptamer and deoxynivalenol, which is applied in the field of biomedicine, can solve the problems of no direct detection method or product of deoxynivalenol, and achieve the effects of ensuring accuracy, improving reliability, and reducing loss

Active Publication Date: 2020-03-17
KUNMING UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Since then, aptamers have been widely used in many aspects such as cell imaging, new drug development, disease treatment and microbial detection, but there is currently no direct detection method or product for DON

Method used

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  • Nucleic acid aptamer specifically bound with vomitoxin, preparation method and application
  • Nucleic acid aptamer specifically bound with vomitoxin, preparation method and application
  • Nucleic acid aptamer specifically bound with vomitoxin, preparation method and application

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Embodiment 1

[0022] 1. Cloning, isolation and sequencing of nucleic acid aptamers and prediction of secondary structure of single-stranded DNA

[0023] Screening: SELEX technology is used to screen out the population of nucleic acid aptamers that can specifically bind to DON through a column that can be coupled to a hydroxyl group.

[0024] Pick a single clone: ​​use the PMD 19-T vector to ligate and transform the PCR product into competent cells, and place the ligated and transformed competent cells on a medium plate with ampicillin by streaking at 37°C overnight.

[0025] Separation: Use the streak method to separate a large number of nucleic acids, and shake the bacteria to obtain the single nucleic acid we need.

[0026] Prediction: Set the temperature to 26°C, Na + The concentration is 150mM, Mg 2+ Concentration of 1mM QGRS mapping was used to predict the secondary structure of the nucleic acid aptamer DON A15 single-stranded DNA molecule specifically binding to vomitoxin. The resu...

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Abstract

The invention belongs to the technical field of biomedicine, and discloses a nucleic acid aptamer specifically bound with vomitoxin, a preparation method and an application. The nucleic acid aptamer is ssDNA and composed of 82 nucleotides, wherein the nucleotide sequence thereof is shown as SEQ ID NO:1. The secondary structure of the nucleic acid aptamer contains protruding rings and stems, wherein DONA15 Gibbs free energy DG is equal to -8.84. The aptamer has a G-quadruplex structure which is more stable. Based on an enzyme-linked oligonucleotide adsorption determination method (ELONA), a series of property evaluations such as specificity, affinity and sensitivity of the nucleic acid aptamer are carried out, and the result shows that DONA15 can be specifically bound with vomitoxin. The nucleic acid aptamer has the characteristics of high specificity and high affinity, and can be applied to subsequent rapid detection of vomitoxin in a test strip.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and in particular relates to a nucleic acid aptamer specifically combined with deoxynivalenol, a preparation method and an application. Background technique [0002] Deoxynivalenol (DON), also known as vomitoxin, is mainly a toxic metabolite of similar chemical structure and biological activity produced by certain species of Fusarium - trichothecene compounds One of. The main producer of DON is Fusarium graminearum, and it has been reported that some other Fusarium can also produce it. DON exists widely in the world, mainly polluting cereal crops such as wheat, barley, and corn, and also contaminating food products. Humans and animals can produce a wide range of toxic effects after eating grains and cereals contaminated by this toxin. In addition, it often contaminates crops together with other mycohalotoxins such as aflatoxin, and can interact with each other after entering the human body....

Claims

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Application Information

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IPC IPC(8): C12N15/115C12N15/10G01N33/569
CPCC12N15/115C12N15/1048G01N33/56961C12N2310/16C12N2310/51C12N2310/531
Inventor 韩芹芹秦鸣蔚夏雪山张晓萌宋玉竹张金阳
Owner KUNMING UNIV OF SCI & TECH
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