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MSO/GO (Mercury-Specific Oligonucleotide/Graphene Oxide) based water environment mercury ion detecting method and kit

A detection method and technology of mercury ions, which are applied in the determination/inspection of microorganisms, biochemical equipment and methods, fluorescence/phosphorescence, etc., can solve the problems of complicated and time-consuming operations, unsatisfactory, expensive costs, etc., and achieve convenient and efficient operation. Simple and low-cost effect

Inactive Publication Date: 2013-06-26
WENZHOU MEDICAL UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Traditional detection methods include atomic absorption spectrophotometry, atomic fluorescence spectrophotometry, high performance liquid chromatography, and mass spectrometry. These detection methods require large-scale instruments, are expensive, complex and time-consuming, and require skilled personnel to operate However, the photochemical analysis technology and electrochemical sensor technology developed in recent years can only perform qualitative or semi-quantitative detection of the tested object due to defects in sensitivity, specificity, or ease of operation, and still cannot meet the current requirements for Hg. 2+ Testing requirements

Method used

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  • MSO/GO (Mercury-Specific Oligonucleotide/Graphene Oxide) based water environment mercury ion detecting method and kit
  • MSO/GO (Mercury-Specific Oligonucleotide/Graphene Oxide) based water environment mercury ion detecting method and kit
  • MSO/GO (Mercury-Specific Oligonucleotide/Graphene Oxide) based water environment mercury ion detecting method and kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Embodiment 1 MSO optimization design

[0039] The MSO sequence selected by the present invention is shown in Table 1, wherein MSO 1 It is a mercury ion-specific oligonucleotide probe (stem-loop structure) reported by Huang Wei et al. In order to improve the sensitivity of mercury ion detection and reduce the cost of DNA synthesis, MSO 1 Carry out an optimized design, remove the non-specific loop part (4bp), intercept the stem part (9bp) at both ends, and mark them as MSO 2 and MSO 3 .

[0040] Table 1 Mercury ion-specific oligonucleotides

[0041]

[0042] Take 2.5 μl MSO (1 μM MSO labeled with FAM fluorescein) 1 , 2μM MSO 2 , 2μM MSO 3 or FAM fluorescein-labeled MSO 2 and unfluorescently labeled MSO 3 Each 1μM mixture), add 5μl 0.5g / L GO, 265μl 20mM metal ion-containing Tris-HCl buffer, react at room temperature for 5 minutes, then add 9.3μM Hg 2+ The solution was reacted for 30 minutes, and the change of the fluorescence intensity at 522 nm was observed. ...

Embodiment 2

[0043] Example 2 Fluorescence-labeled and non-fluorescent-labeled MSO dosage optimization design

[0044] Take 2.5μl of fluorescently labeled MSO at different concentrations (10nM, 100nM, 1μM, 10μM) 2 and unfluorescently labeled MSO 3 Afterwards, 265 μl of 20 mM Tris-HCl buffer solution containing metal ions was added, reacted at room temperature for 5 minutes, and scanned the fluorescence wavelength curve with a fluorescence spectrophotometer. Fluorescence spectrum measurement conditions of the fluorescence spectrophotometer: xenon lamp excitation, slit width 10nm, response time Auto. The excitation wavelength is 496nm, and the scanning range of the emission spectrum is 515-600nm. The measurement is performed at room temperature with a 350μL microcuvette, and the fluorescence value at 522nm is recorded. The fluorescence values ​​at 522nm of different concentrations of fluorescently labeled MSO were recorded for comparison. The result is as figure 1 : When the concentratio...

Embodiment 3

[0045] Example 3 Buffer optimization design

[0046] Prepare Tris-HCl buffer containing metal ions and Tris-HCl buffer without metal ions. Take 2.5 μl of 1 μM fluorescently labeled MSO 2 and unfluorescently labeled MSO 3 Finally, these two buffer solutions were added to scan the fluorescence wavelength curve, and the fluorescence value at the wavelength of 522nm was recorded. Finally, 5 μl of 0.5 g / L GO was added, reacted at room temperature for 5 minutes, and then scanned the fluorescence wavelength to record the change of fluorescence value. figure 2 Among them, when the Tris-HCl buffer containing metal ions was added to the reaction system, the fluorescence quenching efficiency reached (55.191±5.4)% on average before and after adding GO, which was relatively stable; when the Tris-HCl buffer containing no metal ions solution was added to the reaction system, before and after adding GO, the fluorescence quenching efficiency reached (10.108±2.2)% on average, which was much...

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Abstract

The invention relates to an MSO / GO (Mercury-Specific Oligonucleotide / Graphene Oxide) based mercury ion detecting method and a kit for the rapid detection of mercury ion in water. According to the invention, fluorescence-modified and not-fluorescence-modified optimized MSO is adsorbed to the surface of GO by utilizing the acting force of non-covalent bonds (pi-pi superposition); a structure of T-Hg2+-T is formed by the MSO and the mercury ion to ensure that the fluorescently-labeled MSO is released from the surface of the GO and the fluorescence signal is recovered; and the quantitative determination of the mercury ion in solution is realized through detecting the changes of the fluorescence signal in the solution. The method disclosed by the invention has the advantages that the convenient and fast analysis of the mercury ion can be realized without any special condition; the lowest detection limit is 10pM; the sensitivity is greatly increased; the DNA (DeoxyriboNucleic Acid) synthetizing cost is reduced; a solution which is simple, convenient, efficient, sensitive and cost performance is provided for the monitoring of the pollution of the mercury ion in the water; and the method is easy to popularize.

Description

technical field [0001] The invention belongs to the technical field of biological detection, and relates to a method for detecting mercury ions based on mercury-specific oligonucleotide probes (mercury-specific oligonucleotide, MSO) and graphene oxide (Graphene oxide, GO) and a method for detecting mercury in water. A kit for the rapid detection of ions. Background technique [0002] In recent years, with the rapid development of modern industry and agriculture, environmental pollution and poisoning incidents caused by mercury broke out frequently. The discharge of industrial "three wastes", the production of urban and rural garbage, and the unreasonable use of pesticides have made this "invisible killer" threatening our lives and eroding our bodies. Among them, the problem of mercury pollution in my country's water environment is relatively prominent. According to the survey by the Ministry of Agriculture, the area of ​​sewage irrigation areas in my country is about 140×10...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11G01N21/64
Inventor 吕建新吴文鹤
Owner WENZHOU MEDICAL UNIV
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