Method for constructing DNA colorimetric logic gates based on metal ions regulating exonuclease III cleavage

A technology of exonuclease and metal ions, applied in the field of molecular computers, can solve the problems of insufficient sensitive and complete logic system, inapplicability of in-situ detection, etc., and achieve good selectivity

Inactive Publication Date: 2017-09-19
NANCHANG UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the above-mentioned detection methods of metal ions and the construction methods of logic gates require multi-step operations and high-end instruments and equipment, which are not suitable for in-situ detection and are essential for building a sensitive and complete logic system. not enough

Method used

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  • Method for constructing DNA colorimetric logic gates based on metal ions regulating exonuclease III cleavage
  • Method for constructing DNA colorimetric logic gates based on metal ions regulating exonuclease III cleavage
  • Method for constructing DNA colorimetric logic gates based on metal ions regulating exonuclease III cleavage

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Construction of OR logic gates

[0027] The basic principle of DNA logic gate construction is as follows figure 1 shown. Prepare quadruplicate solutions by mixing 20 µL of DNA 1 and 2 hybridization solution with 20 µL of DNA 3 and 4 hybridization solution. Add 20 μL of water to the above solution for (0,0) state, add 10 μL of Ag + (5 μM) is (0,1) state, add 10 μL Hg 2+ (5 μM) is in the (1,0) state, and at the same time add 10 μL Hg 2+ (5 μM) and 10 μL Ag + (5 μM) for the (1,1) state. Add 10 μL of Exo Ш to each sample and incubate at 37 °C for 30 min. Then, 20 μL of signal probe 1 was added to each sample to form G-quadruplexes, and then 10 μL of 2 μM hemin in HEPES buffer solution was added to react at room temperature for 1 h to form G-quadruplexes-heme prime complex. Finally, add 15 µL of TMB-H to each sample 2 o 2 solution, G-quadruplex-heme catalyzed TMB color development, after 4 minutes of reaction, 200 μL of 2 M sulfuric acid was added to terminate the ...

Embodiment 2

[0029] Construction of AND logic gates

[0030] Hybridize 20 μL of DNA 5 and 6 to make quadruplicate solutions. Add 20 μL of water to the above solution for (0,0) state; add 10 μL of Ag + (5 μM) is (0,1) state, add 10 μL Hg 2+ (5 μM) is in the (1,0) state, and at the same time add 10 μL Hg 2+ (5 μM) and 10 μL Ag + (5 μM) for the (1,1) state. Add 10 μL of Exo Ш to each sample and incubate at 37 °C for 30 min. Then, 20 μL of signal probe 1 was added to each sample to form G-quadruplexes, and then 10 μL of 2 μM hemin in HEPES buffer solution was added to react at room temperature for 1 h to form G-quadruplexes-heme prime complex. Finally, add 15 μL of TMB-H to each sample 2 o 2 solution, G-quadruplex-heme catalyzed TMB color development, after 4 minutes of reaction, 200 μL of 2 M sulfuric acid was added to terminate the catalytic reaction. The constructed logic gate was characterized by UV-Vis spectroscopy, the results are as follows image 3 shown. The absorbances of ...

Embodiment 3

[0032] Construction of INHIBIT logic gates

[0033] Mix 20 μL of DNA 1 and 2 hybridization solution with 20 μL of DNA 7 and 8 hybridization solution to make quadruplicate solutions. Add 20 μL of water to the above solution for (0,0) state, add 10 μL of Ag + (5 μM) for (0,1) state, add 10 μL Hg 2+ (5 μM) is in the (1,0) state, and at the same time add 10 μL Hg 2+ (5 μM) and 10 μL Ag + (5 μM) for the (1,1) state. Add 10 μL of Exo Ш to each sample and incubate at 37 °C for 30 min. Then, 20 μL of signal probe 1 was added to each sample to form G-quadruplexes, and then 10 μL of 2 μM hemin in HEPES buffer solution was added to react at room temperature for 1 h to form G-quadruplexes-heme prime complex. Finally, add 15 μL of TMB-H to each sample 2 o 2 solution, G-quadruplex-heme catalyzed TMB color development, after 4 minutes of reaction, 200 μL of 2 M sulfuric acid was added to terminate the catalytic reaction. The constructed logic gate was characterized by ultraviolet-vi...

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Abstract

The invention discloses a DNA colorimetric logic gate construction method based on metal ion regulation and control of exonuclease III shearing action and belongs to the technical field of molecular computer. Double-stranded DNA containing T-T or C-C mismatched base cannot be sheared by exonuclease III. When there exists Hg<2+> or Ag<+>, 3' end completely matched double-stranded DNA formed by action of T-Hg<2+>-T or C-Ag<+>-C can be sheared by exonuclease III, and released single-stranded DNA hybridizes with a DNA signal probe 1 or DNA signal probe 2, thus leading to formation or disintegration of a G-quadruplex structure. Hereby, Hg<2+> and Ag<+> are used as input signals, and logic operation is carried out through the DNA signal probe so as to construct multiple DNA colorimetric logic gates.

Description

technical field [0001] The invention relates to a method for constructing a DNA colorimetric logic gate based on metal ions regulating the shearing action of exonuclease III, and belongs to the technical field of molecular computers. Background technique [0002] Silicon crystal-based computers perform Boolean logic operations by receiving Boolean input signals and generating corresponding electronic output signals. Similarly, molecular computing includes many bottom-up methods to design biomolecules or chemical molecules, and has become another hot research field of computer technology. Researchers have been working on biological and chemical systems that can perform logical operations at the molecular level. DNA logic devices can be constructed by hybridization reactions between nucleic acid molecules, by toehold-controlled strand displacement reactions, by beacon molecular probes, by aptamer binding, by deoxygenation and enzymatic functional reactions construction, so t...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68G06F19/20
CPCC12Q1/6839G16B25/00C12Q2537/119
Inventor 邱建丁郜湾梁汝萍
Owner NANCHANG UNIV
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