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Photoelectrochemical sensor for detecting methylated RNA and detection method thereof

A photoelectrochemical and sensor technology, applied in the direction of material electrochemical variables, scientific instruments, instruments, etc., can solve the problems of low detection sensitivity, cumbersome processing, and low specificity, and achieve high detection sensitivity, simple detection method, The effect of high detection selectivity

Inactive Publication Date: 2019-05-31
SHANDONG AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the above detection techniques basically require 32 P labeling or other radioactive element labeling also requires sophisticated and complex instruments, cumbersome sample pretreatment, professional operators, and low detection sensitivity and specificity.
There is no WS-based 2 A report on the detection of methylated RNA by a photoelectrochemical sensor with Poly(U) polymerase

Method used

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  • Photoelectrochemical sensor for detecting methylated RNA and detection method thereof
  • Photoelectrochemical sensor for detecting methylated RNA and detection method thereof
  • Photoelectrochemical sensor for detecting methylated RNA and detection method thereof

Examples

Experimental program
Comparison scheme
Effect test

preparation example Construction

[0044] (1) Preparation of tungsten sulfide dispersion: Weigh 10-20 mg of tungsten sulfide, add it into deionized water, and ultrasonically disperse for 1-3 hours to prepare a tungsten sulfide dispersion with a concentration of 0.4-5 mg / ml.

[0045] (2) ITO electrode pretreatment: divide the ITO conductive glass into 5×1cm 2 , and then ultrasonically cleaned with ethanol / NaOH mixture (volume ratio 1:1-1:6) for 30-60 minutes, and finally cleaned with acetone and secondary water for 30-60 minutes respectively, and dried at room temperature. stand-by.

[0046] (5) Fixation of tungsten sulfide: Add 25-76 μL of tungsten sulfide dispersion liquid dropwise to the surface of the pretreated ITO electrode, and dry it by infrared light irradiation. Then, wash the electrode with cleaning solution for 3-5 times. Blow dry with nitrogen. The prepared electrode is marked as WS 2 / ITO.

[0047] (4) Modification of nano-gold: Add 16-55 μL nano-gold solution to WS 2 / ITO electrode surface, ...

Embodiment 1

[0060] Example 1: Construction of a photoelectrochemical sensor for detecting methylated RNA

[0061] (1) Preparation of tungsten sulfide dispersion: Weigh tungsten sulfide and add it into deionized water, and ultrasonically disperse for 2 hours to prepare a tungsten sulfide dispersion with a concentration of 2.5 mg / ml.

[0062] (2) ITO electrode pretreatment: divide the ITO conductive glass into 5×1cm 2 , and then ultrasonically cleaned with ethanol / NaOH mixture (volume ratio 1:3) for 60 minutes, and finally cleaned with acetone and secondary water for 30 minutes respectively, and dried at room temperature for use.

[0063] (5) Fixation of tungsten sulfide: 50 μL of tungsten sulfide dispersion was added dropwise to the surface of the pretreated ITO electrode, and dried by infrared light irradiation. Then, wash the electrode with cleaning solution for 3-5 times. Blow dry with nitrogen. The prepared electrode is marked as WS 2 / ITO.

[0064] (4) Modification of nano-gold: ...

Embodiment 2

[0070] Embodiment 2: photoelectrochemical detection

[0071] The electrochemical workstation is used as the signal acquisition instrument, the 500W xenon lamp is used as the visible light source (installed with UV-filtering lens), Ru / U-rich RNA / Ab / MPBA / AuNPs / WS 2 The / ITO electrode is the working electrode, the saturated calomel electrode is the reference electrode, the platinum column electrode is the counter electrode, the 10mM Tris-HCl (pH 7.4) buffer solution containing 0.1M KCl is the detection solution, and the working voltage is -0.3V. Using i-t technology to detect the analyte, establish the relationship between photocurrent and methylated RNA concentration, the linear range is 0.008-30nM, and the calibration curve is I(nA)=-243.898logc-646.944 (R=0.9954) , the detection limit is 2.67pM ( figure 2 ).

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Abstract

The invention discloses a photoelectrochemical sensor for detecting methylated RNA and a detection method thereof. The photoelectrochemical sensor comprises: an electrode, and tungsten sulfide, nanogold, mercaptobenzoic acid, m6A antibody, methylated RNA, poly(U) polymerase mixed with UTP and Ru(NH3)63+ which are modified on the surface of the electrode in order. The photoelectrochemical sensor for detecting methylated RNA and the detection method thereof utilize the good photoelectric activity and biocompatibility of WS2, the good biocompatibility and conductivity of nano gold, the specific recognition performance of antibodies to antigens, employ the Poly(U) polymerase to prolong the RNA sequence and take the Ru(NH3)63+ as a signal amplification unit to achieve the high-sensitivity and high-specificity detection for methylated RNA. The detection method is simple, achieves the miniaturization of the instrument, is easy to operation and achieves the detection for the methylated RNA.

Description

technical field [0001] The invention relates to the technical field of photoelectrochemical analysis, in particular to a photoelectrochemical sensor for detecting methylated RNA and a detection method thereof. Background technique [0002] In the field of epigenetics, RNA methylation is a very important part. Among the many modifications of RNA methylation, N 6 -Methyladenosine (m 6 A) is the most common modification in messenger RNA (mRNA). There is much evidence that m 6 The dynamic regulation of A may have profound effects on the regulation of gene expression. According to recent findings, m 6 A methylation also plays an important and widespread role in various physiological processes such as development, fertility, carcinogenesis, meiosis, and circadian cycle, and is associated with human diseases such as obesity and cancer. Although m 6 The importance of A has been clearly demonstrated, but due to the lack of robust analytical methods, in assessing m 6 Progress ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N27/26G01N27/30G01N27/327
Inventor 殷焕顺王月周云雷艾仕云隋程吉
Owner SHANDONG AGRICULTURAL UNIVERSITY
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