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Target molecule detection method based on nano-Au and nucleic acid structure

A detection method and technology of target molecules, which are applied in the determination/inspection of microorganisms, biochemical equipment and methods, and material analysis by observing the effect on chemical indicators, etc., can solve problems such as unfavorable applications, and achieve the effect of high sensitivity

Inactive Publication Date: 2009-10-21
SHANGHAI INST OF APPLIED PHYSICS - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

These structures are similar to nano-gold but have certain differences. Therefore, one-to-one design is required in the specific operation process, which is not conducive to its application.

Method used

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  • Target molecule detection method based on nano-Au and nucleic acid structure
  • Target molecule detection method based on nano-Au and nucleic acid structure

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] The detection of embodiment 1 thrombin

[0042] Steps: Take 2 μL of thrombin aptamer solution with a concentration of 100 μM and 2 μL of a cDNA solution with a concentration of 100 μM, and add 18 μL of buffer solution (20 mM Tris-acetic acid, pH 7.4, 140 mM NaCl, 1 mM CaCl 2 , 1 mM MgCl 2 ), react at room temperature for 30 min to fully hybridize to form double-stranded probes. Then, 2 μL of an aqueous solution of thrombin with a concentration of 0.1 mM was added to the hybridization solution, and the mixture was reacted for 30 min at room temperature to allow a sufficient reaction. At the same time, a group in which 2 μL of water was added without thrombin was used as a blank experiment. And a group to which 2 μL of 1 mM BSA was added was used as a control. Then, 2 μL of the above reaction solution was added to 100 μL of the gold nano-solution (13 nm, 3.5 nM) prepared above, and the molar number of DNA was controlled to be about 50 times that of the gold nano-partic...

Embodiment 2

[0044] The detection of embodiment 2 lysozyme

[0045] Steps: take 2 μL of lysozyme aptamer solution with a concentration of 100 μM and 2 μL of a cDNA solution with a concentration of 100 μM, add 18 μL of buffer solution (10 mM PB, pH 7.0, 0.2M NaCl), and react at room temperature for 30 minutes to make it Sufficient hybridization is performed to form double-stranded probes. Then, 2 μL of an aqueous solution of lysozyme with a concentration of 0.01-1 mM was added to the hybridization solution, and the mixture was reacted at room temperature for 30 minutes to allow a sufficient reaction. At the same time, a group without adding lysozyme and adding 2 μL of water was used as a blank experiment. And a group to which 2 μL of 1 mM BSA was added was used as a control. Then, 2 μL of the above reaction solution was added to 100 μL of the gold nano-solution (13 nm, 3.5 nM) prepared above, and the molar number of DNA was controlled to be about 50 times that of the gold nano-particles. ...

Embodiment 3 2

[0047] The detection of embodiment 3 divalent mercury ions (one)

[0048] Steps: Take 2 μL of MSO (oligonucleotide specifically binding to mercury ions) solution with a concentration of 100 μM and 2 μL of a cDNA solution with a concentration of 100 μM, respectively, and add 18 μL of buffer solution (10 mM arsinic acid-sodium arsinic acid, pH 6.8, 0.3M NaCl), react at room temperature for 30 min to fully hybridize to form double-stranded probes. Then add 2 μL of 0.01 mM Hg to the hybridization solution 2+ aqueous solution, and reacted at room temperature for 5 minutes to make it fully reacted. without adding Hg 2+ , add 2 μL of water to a group as a blank experiment. In addition, selectivity analysis was carried out through two groups of experiments, one group added 2 μL of 25 mM Ca 2+ , Mg 2+ The other group added 0.5mM mixed ions (Fe 2+ , Cu 2+ ,Co 2+ , Mn 2+ , Ni 2+ , Zn 2+ , Cd 2+ ). After the above reaction solution was diluted 60 times with water, 2 μL was ad...

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Abstract

The invention discloses a target molecule detection method based on nano-Au and nucleic acid structure. The detection method sequentially comprises the following steps: (1) specific DNA which can specifically react with target molecules is sufficiently hybridized with the cDNA of the DNA to form a double-chain capture probe, wherein the target molecules are proteins or ions; (2) target molecule solution is added for full reaction; (3) nano-Au solution with the mol number of 0.01-1 time of that of the specific DNA is added, and the solution is red after reaction; (4) high salting solution with the final concentration of 1-100 mM is added, and the color change of the solution is observed. The detection method has commonality, wide application range, good specificity and high sensitivity, can quickly detect any proteins or any ions with low cost, and does not need DNA mark or dear instruments.

Description

technical field [0001] The invention particularly relates to a target molecule detection method based on nano gold and nucleic acid structure. Background technique [0002] Nucleic acid aptamer (aptamer) refers to the short single-stranded oligonucleotide pairing obtained by screening from a random oligonucleotide library using the SELEX (systematic evolution of ligands by exponential enrichment) in vitro screening technology established at the end of the last century. group, it can specifically bind to the target molecule, thereby undergoing a conformational change itself. [0003] Through in vitro screening technology, nucleic acid aptamers of any substance can theoretically be screened, coupled with the technical characteristics of high-throughput screening and the characteristics of precise recognition of nucleic acid aptamers, easy in vitro synthesis and modification, etc., making nucleic acid aptamers in analytical chemistry and It has broad application prospects in b...

Claims

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Application Information

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IPC IPC(8): G01N21/78C12Q1/68
Inventor 樊春海王丽华宋世平
Owner SHANGHAI INST OF APPLIED PHYSICS - CHINESE ACAD OF SCI
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