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Multi-target quantum-dot mark nucleic acid chip and preparation method and detection method thereof

A nucleic acid chip and quantum dot technology, applied in the field of biosensors, can solve the problems that have not yet been reported on nucleic acid chip research, and achieve the effects of repeated detection synchronization, accurate detection, and simple preparation

Inactive Publication Date: 2009-09-02
THE FIRST AFFILIATED HOSPITAL OF THIRD MILITARY MEDICAL UNIVERSITY OF PLA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Due to these unique quantum advantages, in recent years, the research on the application of quantum dots to FRET has gradually deepened and the scope has been continuously expanded. However, so far, there has been no research report on the use of quantum dots to make nucleic acid chips based on solid supports.

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  • Multi-target quantum-dot mark nucleic acid chip and preparation method and detection method thereof
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  • Multi-target quantum-dot mark nucleic acid chip and preparation method and detection method thereof

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preparation example Construction

[0045] The schematic diagram of the preparation and detection of the first multi-target quantum dot labeled nucleic acid chip is as follows figure 1 As shown, the preparation of the nucleic acid chip is to pass a variety of oligonucleotides without self-complementary sequences and a variety of quantum dots emitting fluorescence of different wavelengths through biotin-avidin (avidin or streptavidin) Coupling is carried out in the system (oligonucleotides of different sequences correspond to quantum dots emitting fluorescence of different wavelengths), and a variety of oligonucleotide probes (QD1-P1, QD2-P2) labeled with quantum dots at one end are prepared; Points and solid supports (for nucleic acid sensors, optional solid supports include silicon wafers, glass slides, tiles, polypropylene membranes or nylon membranes, etc.) by biotin-avidin (avidin or chain Mycoavidin) system is coupled, so that various oligonucleotide probes are fixed on the surface of the solid phase suppor...

Embodiment 1

[0051] 1. Preparation of multi-target quantum dot labeled nucleic acid chip

[0052] 1. Preparation of a variety of quantum dots that emit fluorescence at different wavelengths and are labeled with streptavidin

[0053] (1) Preparation of quantum dots emitting fluorescence of different wavelengths

[0054] Na2 SeSO 3 Solution preparation: In a 250mL three-necked bottle, add Na 2 SO 3 6.302g (0.05mol) and Se powder 1.579g (0.02mol), add high-purity water 100mL to dissolve, stirring, N 2 Under protective conditions, heat up to 90°C for reflux reaction. As the reaction time prolongs, the solution containing gray-black precipitate gradually becomes clear and transparent. After 6 hours of reaction, stop heating and filter with suction to obtain a colorless and transparent filtrate, that is, the concentration is about 0.2mol / L. Na 2 SeSO 3 The solution is stored at a temperature of 70°C (to prevent oxidation) and is used for later use.

[0055] Preparation of NaHSe solution:...

Embodiment 2

[0075] 1. Preparation of multi-target quantum dot labeled nucleic acid chip

[0076] 1. Preparation of a variety of quantum dots that emit fluorescence at different wavelengths and are labeled with streptavidin

[0077] (1) Preparation of various quantum dots emitting fluorescence of different wavelengths

[0078] In a 250mL three-necked bottle, add 100mL of high-purity water, and then add CdCl 2 2.5H 2 O 0.2864g and 222 μL of thioglycolic acid, adjust the pH value of the solution to 11.2 with NaOH with a concentration of 1mol / L, and after 30 minutes of nitrogen gas, add a new anaerobic NaHTe solution under stirring conditions (prepared by the reaction of 80mg of Te powder and 450mg of NaBH ), reflux reaction for 56 hours; take 20mL of reaction solution, add high-purity water to dilute to 100mL, add CdCl 2 2.5H 2 O 0.0576g and 32mL of mercaptoacetic acid, with a concentration of 1mol / L NaOH to adjust the pH value of the solution to 11.2, after nitrogen flow for 30 minutes,...

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Abstract

The invention discloses a multi-target quantum-dot mark nucleic acid chip and a preparation method and a detection method thereof, wherein the nucleic acid chip comprises a solid phase holder and an oligonucleotide probe array fixed on the surface of the solid phase holder, wherein the oligonucleotide probe array comprises at least two oligonucleotide probes which do not contain self-complementary sequences; one end of the oligonucleotide probe marks quantum dots and is fixed by the quantum dots, and the oligonucleotide probes with different sequences are marked by the quantum dots which emit fluorescence with different wavelengths; or the oligonucleotide probe array comprises at least two molecular beacons, wherein one end of the molecular beacon marks quantum dots and is fixed by the quantum dots, the molecular beacons with different sequences are marked by the quantum dots which emit fluorescent with different wavelengths, and the other end of the molecular beacon is marked by a fluorescence quenching group. By utilizing the basic-group complementation pairing principle and the FRET phenomenon, the detection of a plurality of special nucleic acid sequences in a nucleic acid sample to be detected can be simultaneously achieved; in addition, the invention has simple preparation and accurate, sensitive, simple, convenient and rapid detection and can detect a plurality of samples simultaneously.

Description

technical field [0001] The invention relates to a biosensor, in particular to a multi-target quantum dot labeled nucleic acid chip, and also relates to a preparation method and a detection method of the nucleic acid chip. Background technique [0002] In the field of nucleic acid analysis, as an accurate, simple and rapid detection method, nucleic acid sensors have received more and more attention. The basic principle is to immobilize the oligonucleotide probe capable of recognizing a specific nucleic acid sequence on a solid support, perform a hybridization reaction with the nucleic acid sample to be tested during the measurement, and convert the hybridization reaction result into a signal output that can be read. Thereby, the qualitative or quantitative detection of the specific nucleic acid sequence in the nucleic acid sample to be tested can be realized. [0003] The output signal of the nucleic acid sensor during detection can be an electrical signal or an optical sign...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
Inventor 罗阳府伟灵王珏
Owner THE FIRST AFFILIATED HOSPITAL OF THIRD MILITARY MEDICAL UNIVERSITY OF PLA
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