Oligonucleotides chain-based mercury ion fluorescent detection chip, manufacturing method thereof and using method thereof

An oligonucleotide and fluorescence detection technology, applied in the field of biological analysis, can solve the problems of being unsuitable for high-throughput detection and cumbersome operation, and achieve the effects of less reagent consumption, simple production method, and low cost

Active Publication Date: 2011-04-27
SHANGHAI INST OF MICROSYSTEM & INFORMATION TECH CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These methods generally have the characteristics of good selectivity and strong specificity, but the operation is cumbersome and not suitable for high-throughput detection

Method used

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  • Oligonucleotides chain-based mercury ion fluorescent detection chip, manufacturing method thereof and using method thereof
  • Oligonucleotides chain-based mercury ion fluorescent detection chip, manufacturing method thereof and using method thereof
  • Oligonucleotides chain-based mercury ion fluorescent detection chip, manufacturing method thereof and using method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Example 1: Preparation of Hg using Probe A and Probe B 2+ Detection chip.

[0031] Probe A was formulated into a solution with a concentration of 20 μM, then mixed with the same volume of Spotting Solution, and arrayed on the surface of an aldehyde-modified glass slide with the microarray chip production system of Cartesian Company, placed at room temperature, 70% relative Store in humidity for 48-72h for fixation, then immerse the slide in 0.2% SDS at room temperature and shake for a few minutes, then immerse in pure water for a few minutes, then immerse in 0.2% SDS twice, each time for 2min, and then immerse in pure water Twice, 2min each time, let dry. Hybridization solution (10mM MOPS, 100mM NaNO 3 , pH 7.2) Probe B was diluted to a final concentration of 2-5 μM, dropped on the chip, covered with a coverslip, and hybridized at room temperature for 12-16 hours. Then wash with 0.2% SDS, 2×SSC, 0.2×SSC for 3 minutes, and dry it for later use.

Embodiment 2

[0032] Example 2: Using the chips prepared by probe A and probe B to investigate Hg 2+ reaction time kinetics.

[0033] With 10mM MOPS, 100mM NaNO 3 , pH 7.2 diluted to prepare 100μM, 10μM Hg 2+ , the Hg 2+ The solution is added to the prepared chip spots. React at room temperature for 1 hour, take out the chip, and use 10mM MOPS, 100mM NaNO 3 , pH 7.2 buffer washed 3 times and dried. Scan the photos with the chip signal analysis system Scanarray 3000 of General Scanning Company ( figure 2 A-F) and analyze the results ( figure 2 G).

[0034] The results showed that Hg 2+ Inducing the release of probe B is a rapid process, adding 100 μM, 10 μM Hg 2+ After 2 minutes of reaction, compared with the buffer control group, the fluorescence intensity decreased by 63% and 45%, respectively. 95% of the reaction was basically completed in 20 minutes.

Embodiment 3

[0035] Embodiment 3: Utilize the chip prepared by probe A and B to detect different concentrations of Hg 2+ .

[0036] With 10mM MOPS, 100mM NaNO 3 , pH 7.2 diluted to prepare different concentrations of Hg 2+ , plus different concentrations of Hg 2+ The solution was placed on the prepared chip spots at room temperature and reacted for 1 h. With 10mM MOPS, 100mM NaNO 3 , pH 7.2 buffer washed 3 times and dried. Scan the photos with the chip signal analysis system Scanarray 3000 of General Scanning Company ( image 3 A) and analyze the results ( image 3 B).

[0037] The results showed that in Hg 2+ In the presence of ions, the fluorescence intensity at the chip spot weakens, when Hg 2+ When the concentration is 10nM, compared with the fluorescence intensity of the buffer group, the fluorescence intensity at the spot is weakened by 20%, with the increase of Hg 2+ As the concentration increases, the fluorescence signal gradually decreases. Calculated as three times the...

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Abstract

The invention relates to an oligonucleotides chain-based mercury ion fluorescent detection chip, a manufacturing method thereof and a using method thereof. The detection chip can be in convalent binding with a T basic group on a T-enriched oligonucleotides chain specifically through Hg<2+> and mediates T-T pairing of two T-enriched oligonucleotides chains to form a stable intermolecular T-Hg Hg<2+>-T structure and thus induces the release of complementary strands which are crossed with the T-enriched oligonucleotides chain. The use of the detection chip comprises the following steps of: placing a sample to be tested on the chip, keeping a period of time, scanning the chip by using a chip signal analysis system and analyzing a fluorescent signal. The Hg<2+> detection is realized by the change of the fluorescent signal. Higher Hg<2+> concentration in the sample means the fluorescent signal weakens more. The Hg<2+> concentration range capable of being detected by the method is 1nM to 100mu M.

Description

technical field [0001] The invention relates to a mercury ion fluorescence detection chip based on an oligonucleotide chain, a production method and a use method, and belongs to the technical field of biological analysis. Background technique [0002] Mercury is a highly toxic global environmental pollutant, especially its high mobility, persistence, methylation, bioaccumulation, and food chain amplification. The health of animals, plants and humans is also a great threat. The annual emission of mercury in the world is about 15,000 tons, mainly from mercury mines, metallurgy, chlor-alkali industry, electrical industry and combustion of fossil fuels. Mercury exists in various forms in the environment, water-soluble divalent mercury ions (Hg 2+ ) is one of the most common and stable forms of mercury pollution. [0003] How to effectively measure the content of mercury ions in the environment has become a problem facing the majority of analysts. At present, the traditional ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68G01N21/64
Inventor 娄新徽刘美英赵建龙
Owner SHANGHAI INST OF MICROSYSTEM & INFORMATION TECH CHINESE ACAD OF SCI
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