阅读说明：本技术 一种基于核酸染料的电化学检测汞离子的方法 (Method for electrochemically detecting mercury ions based on nucleic acid dye ) 是由 许淑霞 唐刚旭 陈朝霞 于 2019-08-16 设计创作，主要内容包括：一种基于核酸染料的电化学检测汞离子的方法,主要利用金纳米粒子修饰的丝网印刷碳电极作为检测元件,通过稳定的Au-S键的作用在电极表面修饰DNA探针。基于T-T碱基对与Hg<Sup>2+</Sup>特异性结合的特性用以捕捉体系中的Hg<Sup>2+</Sup>,同时引发DNA探针的构性变化。再利用核酸染料GelRed对于双链DNA的嵌入作用促使染料与DNA分子结合,从而引起电极表面的电化学信号升高。基于此,可实现对水样中的Hg<Sup>2+</Sup>无标记检测。该法操作简便,无需标记、成本较低,在实际水样中进行加标检测的结果显示回收率良好,具有良好的应用前景。(A method for electrochemically detecting mercury ions based on nucleic acid dye mainly utilizes a silk-screen printing carbon electrode modified by gold nanoparticles as a detection element, and modifies a DNA probe on the surface of the electrode through the action of a stable Au-S bond. Based on T-T base pairs and Hg 2+ Specific binding property for capturing Hg in system 2+ Simultaneously, the structural change of the DNA probe is triggered. Then the intercalation of the nucleic acid dye GelRed on double-stranded DNA promotes the combination of the dye and DNA molecules, thereby causing the electrochemical signal on the surface of the electrode to be increased. Based on the method, Hg in a water sample can be adjusted 2+ And (4) detecting without marks. The method has simple operation, no need of marking, low cost, and can be used for labeling detection in actual water sampleThe test result shows that the recovery rate is good, and the application prospect is good.)

1. A method for electrochemically detecting mercury ions based on nucleic acid dyes is characterized by comprising the following four steps:

(1) And modifying the gold nanoparticles on the screen-printed carbon electrode by an electrochemical deposition method.

(2) Dripping a DNA probe solution with the concentration of 1 mu M on the surface of the electrode in (1), and incubating at room temperature overnight.

(3) And (3) immersing the electrode in a mercury ion solution with a certain concentration for incubation for 30 minutes.

(4) and (3) immersing the electrode in a GelRed dye solution with the concentration of 5 x, incubating for 10 minutes, detecting an electrochemical signal by a Differential Pulse Voltammetry (DPV) method by using an electrochemical workstation, and analyzing the measurement result.

2. The method of claim 1, wherein the selected DNA probe sequence is 5' -SH- (CH)₂)₆-GGTCTGTCTTCCCCCCCCCCCCTTGTCTGT-3’。

3. The method of claim 1, wherein the electrode is a screen-printed carbon electrode modified by electrochemically deposited gold nanoparticles.

4. The method of claim 1, wherein the dye used to enhance the electrochemical signal is GelRed.

5. The method of claim 1, wherein the Tris-HCl (Tris-hydroxymethyl-aminomethane-hydrochloric acid) buffer solution has a pH of 7.4.

Technical Field

The invention relates to quantitative detection of mercury ions by using an electrochemical biosensor, and belongs to the technical field of biosensing.

Background

With the development of human society and industry, the problem of environmental pollution is increasingly highlighted, and the survival and development of human beings are seriously threatened. Heavy metal pollution refers to environmental pollution caused by heavy metals and their compounds, mainly caused by artificial factors such as mining, industrial waste gas and water discharge, etc. Wherein the mercury ion (Hg)²⁺) Is a water-soluble and high-risk heavy metal ion, can not only destroy the ecological environment, but also cause no harm to aquatic organisms and human healthDamage can be reversed. Hg is a mercury vapor²⁺It can seriously damage various organs of the human body, including the brain, kidneys, central nervous system and digestive system. Hg in drinking water according to the United states Environmental Protection Agency (EPA) regulations²⁺Should not exceed 10 nM. Conventional Hg²⁺The analysis method includes Atomic Absorption Spectroscopy (AAS), Atomic Fluorescence Spectroscopy (AFS), High Performance Liquid Chromatography (HPLC), inductively coupled plasma mass spectrometry (ICP-MS), and the like. These highly sensitive, highly accurate and highly accurate analytical methods have been widely used for Hg in environmental and biological samples²⁺Detection of (3). However, these analytical methods have some disadvantages such as the need for expensive equipment and complicated operating procedures. Furthermore, these techniques cannot be used for field testing due to the large scale instrumentation required. In comparison, the electrochemical analysis method has the advantages of simple equipment and process, high sensitivity, quick analysis and the like.

In recent years, biosensors based on DNA molecules that specifically recognize a target substance have been vigorously developed because they exhibit advantages such as rapidity, high sensitivity, and high selectivity. Ono and Togashi discovered Hg in 2004²⁺And specific mismatch between T-T base pairs in DNA molecules, for the detection of Hg by DNA biosensors²⁺A basis is provided.

Disclosure of Invention

The invention aims to construct a label-free electrochemical biosensor based on nucleic acid dye, which is used for quickly and sensitively detecting mercury ions.

The gold nanoparticles are modified on the surface of the disposable silk-screen printing carbon electrode by using an electrochemical deposition technology, so that the purposes of improving the conductivity of the silk-screen printing carbon electrode and facilitating modification of a DNA probe are achieved. Then modifying the DNA probe with one end modified with-SH to the surface of the electrode through the action of Au-S bond to capture Hg in the solution²⁺The purpose of enhancing electrochemical signals is achieved by virtue of the intercalation effect of a nucleic acid dye GelRed and a DNA double strand. Therefore, the mercury ion detection device can achieve high selectivity, no mark and high sensitivity detection of mercury ions in water.

The technical scheme of the invention is as follows:

(1) And modifying the gold nanoparticles on the screen-printed carbon electrode by an electrochemical deposition method.

(2) dripping a DNA probe solution with the concentration of 1 mu M on the surface of the electrode in (1), and incubating at room temperature overnight.

(3) Immersing the electrode in (2) into Hg with a certain concentration²⁺Incubate in solution for 30 minutes.

(4) And (3) immersing the electrode in a GelRed dye solution with the concentration of 5 x, incubating for 10 minutes, detecting an electrochemical signal by a Differential Pulse Voltammetry (DPV) method by using an electrochemical workstation, and analyzing the measurement result.

Effects of the invention

Compared with the prior art, the invention has the following advantages:

(1) The sensitivity is high. The method can detect nM Hg²⁺Can reach Hg in the actual water environment²⁺And (5) content standard.

(2) The cost is low. The method only needs a single DNA probe molecule and does not need to be marked, thereby greatly reducing the detection cost.

(3) The operation is simple. The detection of the sample can be completed in a short time without other special sample pretreatment processes.

Detailed Description