DNA tetrahedral nano-structure signal probe and application thereof

A technology of signaling probes and nanostructures, applied in the biological field, can solve the problems of low efficiency of enzyme reactions

Inactive Publication Date: 2015-05-27
SHANGHAI ADVANCED RES INST CHINESE ACADEMY OF SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, since a single-stranded DNA signaling probe usually only binds to one HRP enzyme, the efficiency of the enzymatic reaction is not high

Method used

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  • DNA tetrahedral nano-structure signal probe and application thereof
  • DNA tetrahedral nano-structure signal probe and application thereof
  • DNA tetrahedral nano-structure signal probe and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0081] Four single-stranded DNAs used to assemble tetrahedral DNA nanostructure signal probes: tetra-a-reporter (77bp, molecular weight 23718.43, ssDNA); tetra-b-biotin (55bp, molecular weight 17032.04, 5' end modified biotin ssDNA ); tetra-c-biotin (55bp, molecular weight 16909.915' end modified biotin ssDNA); tetra-d-biotin (55bp, molecular weight 16888.84, 5' end modified biotin); all were synthesized by Shanghai Shenggong Company.

[0082] The four DNAs that constitute the tetrahedral nanostructure signal probe contain three structural domains, each of which is complementary to the corresponding structural domains of the other three single-stranded DNAs (17 pairs of bases are complementary), and each single-stranded DNA surrounds the four sides One face of the tetrahedron is a circle, and each vertex contains two bases (non-complementary, flexible) for bending, and the 3' and 5' ends of the single-stranded DNA converge on the four vertices of the tetrahedron. Tetra-a-repor...

Embodiment 2

[0097] The electrochemical detection method is as follows:

[0098] 1) Self-assembled tetrahedral DNA capture probe and tetrahedral DNA nanostructure signal probe

[0099] Synthesis of capture probes: take equal amounts of four single-stranded DNAs of tetra-a, b, c, and d, and use TM buffer (20mM tris, 50mM MgCl 2 ,pH8.0) diluted to a concentration of 1uM and a volume of 100uL. After 95 degrees Celsius for 3 minutes, immediately cool down to 4 degrees Celsius.

[0100] Synthesis of signal probes: take equal amounts of four single-stranded DNAs of tetra-a-reporter, b-biotin, c-biotin, and d-biotin, and use TM buffer (20mM tris, 50mM MgCl 2 ,pH8.0) diluted to a concentration of 1uM and a volume of 100uL. After 95 degrees Celsius for 3 minutes, immediately cool down to 4 degrees Celsius.

[0101] 2) Clean and polish the electrodes and assemble

[0102] Take a gold electrode with a diameter of 2mm, first polish it with 0.3um and 0.5um alumina powder in sequence, then use etha...

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Abstract

The invention provides a DNA tetrahedral nano-structure signal probe. The signal probe is a DNA signal probe having a tetrahedral base, wherein the signal probe is formed by self-assembling single-stranded DNA of a DNA recognition sequence extending from 5' end and single-stranded DNA of three chains with 5' ends modified with labeled molecule; the DNA recognition sequence extends from a vertex of the tetrahedral structure, the DNA recognition sequence is complementarily combined with to-be-detected target DNA or miRNA, and other three vertexes, due to the labeled molecules, are specifically combined with modified signaling modules, so as to achieve multi-signal amplification. The signal probe disclosed by the invention greatly improves the sensitivity of the target DNA or miRNA subjected to electrochemical detection, and a limit of detection can be as low as 1fM.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a DNA tetrahedron nanostructure signal probe and its application. Background technique [0002] DNA electrochemical biosensors have great potential applications in various fields such as clinical medicine, food inspection, environmental detection and anti-terrorism due to their advantages of rapidity, sensitivity, low cost and easy miniaturization. An important aspect that determines the performance of DNA electrochemical biosensors is the signal amplification system. [0003] As a biomarker, DNA plays a very important role in disease diagnosis. Therefore, ultrasensitive DNA sensors for detecting biomarkers at low concentrations of DNA in clinical samples are of great importance. Among them, the electrochemical DNA sensor is a promising one because of its fast response, ease of use, low cost, and direct DNA detection. Electrochemical DNA sensors were first invented in 2003, among ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11G01N27/327G01N27/48
Inventor 宓现强曾冬冬
Owner SHANGHAI ADVANCED RES INST CHINESE ACADEMY OF SCI
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