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Long afterglow molecular beacon probe as well as construction method and application thereof

A technology of molecular beacon probes and molecular beacons, applied in the field of time-resolved molecular beacon probes and their construction, and molecular beacon probes, can solve the problems of complex and cumbersome synthesis steps, unfavorable construction of molecular beacons, and reduced sensitivity , to achieve the effects of good water solubility, stable luminescence and long emission life

Active Publication Date: 2021-04-30
CIXI INST OF BIOMEDICAL ENG NINGBO INST OF MATERIALS TECH & ENG CHINESE ACAD OF SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] The above existing technologies mainly have the following problems: 1. When organic dye molecules are used as fluorescent donors to construct molecular beacons, their visible light excitation characteristics will cause optical interference such as matrix background fluorescence and scattered light, resulting in reduced sensitivity; Rare earth Eu 3+ The synthesis steps of complexes are complex and cumbersome, and their water solubility is usually poor, which is not conducive to the construction of molecular beacons

Method used

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  • Long afterglow molecular beacon probe as well as construction method and application thereof
  • Long afterglow molecular beacon probe as well as construction method and application thereof
  • Long afterglow molecular beacon probe as well as construction method and application thereof

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preparation example Construction

[0056] In some embodiments, the fluorescent donor material is polyacrylic acid modified ZnGa 2 o 4 :Cr 3+ long-lasting nanocrystals, and, the polyacrylic acid-modified ZnGa 2 o 4 :Cr 3+ The preparation method of long afterglow nanocrystals comprises:

[0057] Evenly mix the alkaline substance, oleic acid, water and ethanol to form the first mixed solution;

[0058] The Zn source, the Ga source, and the Cr source are uniformly mixed according to a molar ratio of 1:2:0.01-0.3 to form a second mixed solution;

[0059] Add the second mixed solution dropwise to the first mixed solution, and uniformly stir at room temperature for 10-60 minutes, and then conduct a hydrothermal reaction at 180-220° C. for 8-24 hours to obtain ZnGa 2 o 4 :Cr 3+ long-lasting nanocrystals; and,

[0060] Using polyacrylic acid for the ZnGa 2 o 4 :Cr 3+ modified with long-lasting nanocrystals to obtain the polyacrylic acid-modified ZnGa 2 o 4 :Cr 3+ Long persistence nanocrystals.

[0061] F...

Embodiment 1

[0102] Example 1: Construction of near-infrared long-lasting molecular beacon probe and application of miRNA21 detection

[0103] (1) Preparation of near-infrared long-lasting luminescent materials and surface water-soluble modification

[0104] Add 0.3g of NaOH to a mixed solution of 3mL of oleic acid, 4mL of deionized water and 10mL of absolute ethanol, stir for 15min until NaOH is completely dissolved; then add 0.5mL of Zn(NO 3 ) 2 (1M), 2mLGa(NO 3 ) 3 (2M), and 40μL Cr(NO 3 ) 3 (0.05M) were mixed, and stirred uniformly at room temperature for 30 minutes; then the mixed solution was transferred to a 25mL hydrothermal kettle (lined with polytetrafluoroethylene), and heated in an oven at 210°C for 16 hours. After naturally cooling to room temperature, the reaction solution in the hydrothermal kettle was centrifuged to obtain the product, and the precipitate was washed three times with a mixed solvent of cyclohexane and ethanol. Finally, the precipitate was dried in a va...

Embodiment 2

[0107] Example 2: Construction of near-infrared long-lasting nucleic acid aptamer molecular beacon probe and application of thrombin detection

[0108] (1) Preparation of near-infrared long-lasting luminescent materials and surface water-soluble modification

[0109] Add 0.3g of NaOH to a mixed solution of 3mL of oleic acid, 4mL of deionized water and 10mL of absolute ethanol, stir for 15min until NaOH is completely dissolved; then add 0.5mL of Zn(NO 3 ) 2 (1M), 2mL Ga(NO 3 ) 3 (2M), and 8μL (Cr(NO 3 ) 3 (0.01M) were mixed and stirred uniformly at room temperature for 20 minutes; then the mixed solution was transferred to a 25mL hydrothermal kettle (lined with polytetrafluoroethylene), and heated in an oven at 220°C for 8 hours. After naturally cooling to room temperature, the reaction solution in the hydrothermal kettle was centrifuged to obtain the product, and the precipitate was washed three times with a mixed solvent of cyclohexane and ethanol. Finally, the precipit...

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Abstract

The invention discloses a long afterglow molecular beacon probe as well as a construction method and an application thereof. The long afterglow molecular beacon probe comprises: a molecular beacon; and a fluorescence donor material and an energy acceptor material respectively modified at two tail ends of the molecular beacon; wherein the fluorescence donor material comprises a rare earth-based long afterglow luminescent material, phosphorescence / long afterglow luminescent carbon quantum dots, phosphorescence semiconductor quantum dots and the like, the energy acceptor material is a quenching agent, and the quenching agent comprises an optical material with a long afterglow quenching performance. The long-afterglow luminescent material adopted by the invention is simpler in preparation steps, good in water solubility, more stable in luminescence and longer in emission life, and the constructed time-resolved molecular beacon probe based on long-afterglow luminescence is higher in signal-to-noise ratio and sensitivity, and is applied to high-sensitivity detection of micro RNA in an in-vitro biological sample; and a new strategy is provided for the design of the molecular beacon probe.

Description

technical field [0001] The present invention relates to a molecular beacon probe, in particular to a time-resolved molecular beacon probe based on rare earth long-lasting luminescence and its construction method, which is applied to the highly sensitive detection of microRNA or other nucleic acids in biological samples in vitro detection, and belongs to the technical field of nano-fluorescence detection. Background technique [0002] Molecular beacon (MB) is a fluorescently labeled oligonucleotide chain, generally composed of three parts: ①Ring region: composed of 15-30 nucleotides that can specifically bind to the target molecule;② Stem region: generally composed of 5 to 8 base pairs that can undergo reversible dissociation; ③ Fluorescence donor and acceptor: the two ends of the molecular beacon are labeled with fluorescent dye and quencher dye respectively. When there is no target molecule, the fluorescence donor and the quencher group of the molecular beacon are very clo...

Claims

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Application Information

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IPC IPC(8): C12Q1/6818C12Q1/6806C12N15/11C12N15/115G01N33/532G01N33/573G01N33/68
CPCC12Q1/6818C12Q1/6806C12N15/115G01N33/573G01N33/68G01N33/532G01N2333/974C12N2310/16C12N2310/531C12Q2563/103C12Q2565/1015
Inventor 王宇辉郑建萍王秀华
Owner CIXI INST OF BIOMEDICAL ENG NINGBO INST OF MATERIALS TECH & ENG CHINESE ACAD OF SCI
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