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Aptamer for specifically recognizing diethylstilbestrol

A nucleic acid aptamer, diethylstilbestrol technology, applied in biochemical equipment and methods, instruments, analytical materials, etc., can solve the problems of high cost, high detection cost, cumbersome and time-consuming biological antibody preparation process, and achieve low cost and application. promising effect

Inactive Publication Date: 2018-11-06
浙江省产品质量安全检测研究院
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  • Abstract
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AI Technical Summary

Problems solved by technology

The first two methods have high sensitivity and accuracy, but require expensive precision instruments and well-trained professional technicians, the detection cost is relatively high, and are suitable for confirmation detection in large laboratories; ELISA is easy to operate and suitable for a large number of samples Rapid screening for detection, but the preparation process of biological antibodies is cumbersome, time-consuming and expensive

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  • Aptamer for specifically recognizing diethylstilbestrol
  • Aptamer for specifically recognizing diethylstilbestrol

Examples

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Embodiment 1

[0018] Example 1: Index enrichment screening of diethylstilbestrol aptamers by means of streptavidin magnetic beads

[0019] 1. Random nucleic acid library and primers (completed by Sangon Biosynthesis (Shanghai) Co., Ltd.), the nucleotide sequence is as follows:

[0020] Random nucleic acid library: 5'-ACGGACTGAGCTCGAGTCT-(N40)-AGATTGACCGACGGATGCG-3'

[0021] Among them, N40 represents 40 random nucleotides.

[0022] Upstream primer P1: 5'- ACGGACTGAGCTCGAGTCT -3'

[0023] Downstream primer P2: 5'-CGCATCCGTCGGTCAATCT-3'

[0024] 5'Biotinylation downstream primer Biotin-P2: 5'-Biotin-CGCATCCGTCGGTCAATCT -3'

[0025] 2. PCR amplification conditions and preparation conditions of single-strand sub-library

[0026] Using the random nucleic acid library as a template, carry out asymmetric PCR amplification, as follows:

[0027] (1) The PCR reaction system is:

[0028] Template 23 μL, upstream primer 2 μL, downstream primer 1 μL, dNTP (2.5mM each) 4 μL, 10×PCR amplification bu...

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Abstract

The invention belongs to the technical field of biological detection and in particular relates to an aptamer for specifically recognizing diethylstilbestrol. The preparation method comprises the following steps: performing diethylstilbestrol binding screening by utilizing a magnetic bead method by virtue of systematic evolution of ligands by exponential enrichment; performing multi-round enrichment screening on nucleotide sequences subjected to PCR (Polymerase Chain Reaction) amplification binding, thereby obtaining the high-affinity aptamer capable of specifically recognizing diethylstilbestrol. The aptamer provided by the invention can be synthesized by commercial corporations, can be artificially synthesized and labeled, and is low in cost; the aptamer provided by the invention can be specifically bound to the diethylstilbestrol, a rapid detection method of the diethylstilbestrol in foods or environments can be established, and the application prospects are wide.

Description

technical field [0001] The invention belongs to the technical field of biological detection, in particular to a nucleic acid aptamer that specifically recognizes diethylstilbestrol. Background technique [0002] As an environmental hormone closely related to food safety risks, diethylstilbestrol has the effect similar to estrogen. Even in a very small amount, it may lead to abnormal behavior of animals and humans, decreased reproductive ability, precocious puberty and other diseases. Since diethylstilbestrol can promote animal growth and increase the lean meat rate, it is often illegally used in animal feeding, causing drug residues in animal viscera, muscle, and even eggs and milk, causing food safety hazards. [0003] The determination methods of diethylstilbestrol mainly include high performance liquid chromatography-tandem mass spectrometry, high performance liquid chromatography and enzyme-linked immunoassay. The first two methods have high sensitivity and accuracy, bu...

Claims

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Application Information

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IPC IPC(8): C12N15/115G01N33/533G01N33/532
CPCG01N33/532G01N33/533C12N15/115C12N2310/16
Inventor 张慧姜侃陈小珍张东雷
Owner 浙江省产品质量安全检测研究院