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Nanogold dual-probe system based on DNA-encoded cycle program

A coding loop and dual probe technology, applied in the field of loop programming detection technology, can solve the problems of low imaging sensitivity and unsatisfactory detection effect, and achieve the effect of improving sensitivity

Active Publication Date: 2022-02-11
EAST CHINA UNIV OF SCI & TECH
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Problems solved by technology

[0004] Although these gold nanoparticle-based nanoprobes can be used for intracellular imaging, when the analytes are miRNAs with very low abundance in cells, the imaging sensitivity is low and the detection effect is not ideal.

Method used

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  • Nanogold dual-probe system based on DNA-encoded cycle program
  • Nanogold dual-probe system based on DNA-encoded cycle program

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Embodiment 1

[0025] (1) According to the existing literature, a one-step method was used to synthesize 60nm gold nanoparticles. 4.8mL of HAuCl 4 (1%) was added into a round-bottomed flask containing 100mL of water and heated to boiling, and 10mL solution of 0.1g sodium citrate was added dropwise and continued to boil for 15min. The solution presented a reddish brown color to prepare gold nanoparticles of about 60nm.

[0026] (2) The two DNAs were first thawed at room temperature for about 10 minutes, and centrifuged at 3000 rpm for 5 minutes. According to the instructions, the solution was dissolved in PBS (pH=7.4) to obtain a 10 μM DNA solution, and the 60 nm gold solution was centrifuged at 5800 rpm for 5 min. After aspirating the supernatant, add 1 mL of deionized water. Take 10 μL of 10 μM DNA solution in 1 mL of 60 nm gold nano solution, and stabilize it in a shaker at 25° C. and 120 rpm for 24 hours. Then centrifuge at 5800rpm for 5min, take the precipitate and add deionized water...

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Abstract

The present invention provides a nano-gold dual-probe system based on a DNA coding cycle program, including a seed probe and a fuel probe, a base sequence on the seed probe matches the miRNA base sequence, and the fuel probe and the miRNA base sequence match at the same time. The base pairs on the seed probes are completely matched, and the dual-probe system can realize the circular detection of miRNA in tumor cells. This method provides a new fluorescence detection method, which improves the sensitivity of the detection method by realizing the circular detection of intracellular miRNA through seed probes and fuel probes, and realizes ultrafast and ultrasensitive imaging detection of intracellular miRNA, realizing Early detection and prevention of tumor cells. The seed probes and fuel probes designed by the present invention can detect the circulation of a small amount of miRNA in cells, and provide a new method for studying the intracellular physiological processes that they participate in.

Description

technical field [0001] The invention relates to the fields of cell imaging technology, cell pathway regulation and nanometer material preparation. Specifically, it is a cycle programming detection technology for microRNA (miRNA) in living cells based on dark field / fluorescence microscopy imaging technology. Background technique [0002] In recent years, the functions and importance of miRNAs in cellular regulation have attracted increasing attention. Recent research investigations have shown that the overexpression of miRNA-21 is closely related to the occurrence of various cancers, and it is considered to be the most reflective marker in cancer cells and used for cancer diagnosis and treatment. Therefore, the development of effective and sensitive miRNA live cell imaging detection technology is of great significance for the elucidation of pathophysiological mechanisms. [0003] Fluorescence spectroscopy is a single-molecule detection method with high sensitivity. The main...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/64G01N21/73C12Q1/6886
CPCG01N21/6458G01N21/6486G01N21/73C12Q1/6886
Inventor 龙亿涛钱若灿吕键
Owner EAST CHINA UNIV OF SCI & TECH
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