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A fluorescent method for precise quantification of structural integrity of DNA origami

A technology of origami structure and integrity, applied in the field of nano-biological materials and biological detection, can solve the problems of being easily affected by the environment, structural stability, structural changes, etc., and achieve the effect of accurate quantitative methods and easy evaluation operations

Active Publication Date: 2020-06-02
SHANGHAI NAT ENG RES CENT FORNANOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] However, DNA materials are more susceptible to the influence of the environment than inorganic materials, especially when applied to the human body, they are susceptible to structural changes due to interference from temperature, ion concentration, and other impurities, resulting in changes in their own excellent properties.
At present, there have been many research results in this field, such as the DNA origami structure can only maintain a stable structure for about 20 hours in serum; the rise of temperature will cause its structure to dissociate; the increase or decrease of ion concentration will cause structural changes ; Exposure to ultraviolet light or ozone environment will also affect its structural stability
So how much does environmental change affect structural stability? There are many qualitative studies, but there is no quantitative method.

Method used

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  • A fluorescent method for precise quantification of structural integrity of DNA origami
  • A fluorescent method for precise quantification of structural integrity of DNA origami

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Experimental program
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Effect test

Embodiment 1

[0025] The DNA triangular origami structure synthesized above was purified in an ultrafiltration centrifuge tube at a speed of 3000 g for 10 minutes, and centrifuged 3 times to remove excess staple chains.

[0026] (1) Set the final concentration of the purified DNA triangula to 2nM, take 10 μl of the DNA triangula solution, add 5 μl of 20×Eva Green, and add 1×TAE-Mg 2+ 85 μl of buffer solution, measure its fluorescence intensity, the structure remains intact (0%);

[0027] (2) Control Mg 2+ Concentration is 1 / 10 in the original buffer solution, stand for 5 minutes, measure its fluorescence intensity, the structure changes by 28.36%;

[0028] (3) Restore the Mg in the solution 2+ Concentration is 1×TAE-Mg 2+ , the fluorescence intensity was measured again, and the structure recovered to 0.88%, with figure 2 a.

[0029] Always keep the concentration of DNA triangular origami at 0.2 nM. Determine the completeness of the DNA origami structure according to the size of the f...

Embodiment 2

[0031] The DNA triangular origami structure synthesized above was purified in an ultrafiltration centrifuge tube at a speed of 3000 g for 10 minutes, and centrifuged 3 times to remove excess staple chains.

[0032] (1) Set the final concentration of the purified DNA triangula to 2nM, take 10 μl of the DNA triangula solution, add 5 μl of 20×Eva Green, and add 1×TAE-Mg 2+ 85 μl of buffer solution, measure its fluorescence intensity;

[0033] (2) Control Mg 2+ The concentration is 1 / 10 of the original buffer solution, let it stand for 1 hour, and measure its fluorescence intensity, the structure has changed by 33.45%;

[0034] (3) Restore the Mg in the solution 2+ Concentration is 1×TAE-Mg 2+ , measured its fluorescence intensity again, the structure could not be recovered, and the disturbance of the concentration of the recovery solution caused a greater damage to the structure to 37.91%. figure 2 b.

[0035] Always keep the concentration of DNA triangular origami at 0.2 n...

Embodiment 3

[0037] The DNA triangular origami structure synthesized above was purified in an ultrafiltration centrifuge tube at a speed of 3000 g for 10 minutes, and centrifuged 3 times to remove excess staple chains.

[0038] (1) Set the final concentration of the purified DNA triangula to 2nM, take 10 μl of the DNA triangula solution, add 5 μl of 20×Eva Green, and add 1×TAE-Mg 2+ 85 μl of buffer solution, measure its fluorescence intensity;

[0039] (2) Control Mg 2+ The concentration is 1 / 10 of the original buffer solution, let it stand for 5 minutes, measure its fluorescence intensity, and the structure changes by 33.03%;

[0040] (3) Add a small amount of Q water (10 μl), observe the effect of solution disturbance on the fluorescence intensity, and measure the fluorescence intensity again, the structure changes by 32.17%, attached figure 2 left picture.

[0041] Always keep the concentration of DNA triangular origami at 0.2 nM. Determine the completeness of the DNA origami struc...

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Abstract

The invention relates to a fluorescence precise quantification method of the integrity of a DNA origami structure. The method includes the steps of synthesizing a DNA triangular origami structure andquantifying the change of the DNA triangular origami structure along with the change of ion concentration through a fluorescence spectrum. Specifically, an EvaGreen fluorescent dye serves as the quantification agent, and the integrity of the DNA triangular origami structure during environment changes is precisely quantified through fluorescence intensity. The invention belongs to the field of nanometer biological materials and biological detection. The method has the advantages of being precise, simple, simple in evaluating operation and capable of meeting the application requirements of the DNA nanometer origami structure in future.

Description

technical field [0001] The invention relates to a method for accurately quantifying the structural integrity of DNA origami with fluorescence, in particular to using Eva Green fluorescent dye as a quantitative reagent to accurately quantify the structural integrity of the DNA triangular origami structure when the environment changes through fluorescence intensity. The invention belongs to the field of nano biological material and biological detection. Background technique [0002] DNA origami structure has received extensive attention due to its precise and controllable structure, easy modification, and good biocompatibility. In recent years, studies have found that the DNA double helix has special properties such as electrical conductivity and pH responsiveness. The DNA double helix can control the structural size accuracy to the order of angstroms through the principle of complementary base pairing (10 -10 ), its 3' and 5' ends are easy to modify various chemical groups, ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/64
CPCG01N21/6428G01N2021/6439
Inventor 何丹农王萍陈益徐艳叶恺金彩虹
Owner SHANGHAI NAT ENG RES CENT FORNANOTECH
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