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Method of regulating and controlling in-site catalytic polymerization reaction with bioenzyme on nano material on the basis of DNA

A technology for catalytic polymerization and nanomaterials, applied in the field of fixed-site catalytic polymerization, can solve the problems of no electroactivity, difficult to control the enzymatic polymerization reaction at a fixed point, hinder the formation of ordered polymers, etc., to achieve rapid reaction, low toxicity, biological The effect of high safety

Inactive Publication Date: 2017-07-14
SHANGHAI NAT ENG RES CENT FORNANOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, enzymatic polymerization is usually accompanied by side reactions, and the disordered polymer structure not only has no electrical activity, but also hinders the formation of ordered polymers.
And for nano-scale materials, it is difficult to achieve fixed-point control of enzymatic polymerization.

Method used

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  • Method of regulating and controlling in-site catalytic polymerization reaction with bioenzyme on nano material on the basis of DNA
  • Method of regulating and controlling in-site catalytic polymerization reaction with bioenzyme on nano material on the basis of DNA
  • Method of regulating and controlling in-site catalytic polymerization reaction with bioenzyme on nano material on the basis of DNA

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Mix 400 μL of 50 nm spherical gold nanoparticles (Au NPs, 1 nM) with single-stranded thiol-modified DNA 1 at a ratio of 1:3000, and slowly add sodium chloride (NaCl) and phosphate buffer (PB) to make Their final concentrations reached 0.1 M and 10 mM, respectively. After the mixture was incubated at 25°C for 8 hr, excess DNA was washed by centrifugation three times with 10 mM phosphate buffered saline (PBS). The resulting pellet was resuspended in PBS, biotin-modified single-stranded DNA 2 was added at a ratio of 1:3000, and incubated at 25°C to hybridize DNA 1 and DNA 2. After reacting for 8 hrs, centrifuge three times and use PBS to wash off excess DNA 2. After the obtained precipitate is resuspended in PBS, add 2 μL of 1 mM avidin-HRP to incubate for 30 min, centrifuge three times to wash off excess avidin-HRP, and use 400 Resuspend the pellet in μLPBS. Take 50 μL of the 200-fold diluted resuspension solution and add it dropwise on the cleaned conductive glass slid...

Embodiment 2

[0032] Design the DNA origami structure (the original sequence of the origami staple chain is the DNA sequence 14-221), DNA origami B-56 ​​position protrudes a single strand-DNA hybridizes with the sulfhydryl-modified complementary strand DNA 3, at A-28, B-28, C-28 (respectively DNA 4, 5, 6) positions protrude single-stranded DNA. Mix 2 nM long-strand M13 DNA with 20 nM staple-strand DNA (including the replacement strand) in 1×TAE-Mg 2+ Mix well in buffer (40 mM tris(hydroxymethyl)aminomethane, 2 mM acetic acid, 2 mM disodium edetate, 12.5 mM magnesium acetate, pH 8.0), incubate at 95°C for 5 min, and slowly anneal to Centrifuge three times at 25°C with a 100 kd ultrafiltration tube to filter out excess single strands, recover the triangular origami structure in the tube, and quantify its concentration by ultraviolet light. HRP and SPDP were incubated at a ratio of 1:20 for 2 hr (PBS, pH=8.5), and ultrafiltered three times to wash away excess SPDP. Subsequently, HRP was incu...

Embodiment 3

[0035] Mix rod-shaped gold nanoparticles (Au NRs) and sulfhydryl-modified DNA 7 at a ratio of 1:3000, and slowly add sodium chloride (NaCl) and phosphate buffer (PB) to make the final concentrations of 0.1 M and 10 M, respectively. mM. After the mixture was incubated at 25°C for 8 hr, excess DNA was washed by centrifugation three times with 10 mM PBS. Design the DNA origami structure (the original sequence of the origami staple chain is DNA sequence 14-221), DNA origami B-56 ​​position protrudes a single-stranded DNA to hybridize with the biotin-modified complementary strand DNA 5, and at the same time at B-45 , B-37, B-08, B-16, B-23 (DNA 8, 9, 10, 11, 12, respectively) stretch out the arm strands to capture Au NRs modified with DNA 4. Mix 2 nM long-strand M13 DNA with 20 nM staple-strand DNA in 1×TAE-Mg 2+Mix well in the buffer, incubate at 95°C for 5 minutes, anneal to 25°C, and filter out excess single strands with a 100 kd ultrafiltration tube. 5 nM of the obtained DNA...

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Abstract

The invention relates to a method of regulating and controlling an in-site catalytic polymerization reaction with a bioenzyme on a nano material on the basis of DNA. According to different adsorption capabilities of a polymerization substrate to single- and double-chain DNAs, single- and double-chain DNAs are stretched out at special sites of a nano structure; the bioenzyme is then conjugated to the nano material to perform the catalytic polymerization reaction; a reagent, which is required in the bioenzyme catalytic polymerization reaction, is added to the nano material after the bioenzyme is conjugated thereto, and according to different demands, reaction conditions are controlled so as to in-site catalyze generation of a high-molecular polymer on the single- and double-chain DNAs on the original nano structure. In the method, by controlling the reaction conditions, nano structures, such as gold nano particle, DNA origami and the like, are modified with the bioenzyme, thereby catalytically generating the polymer on the material modified with the single- and double-chain DNAs by means of the bioenzyme. The method can quickly, on special sites, catalyze an ordered polymerization reaction and can be used for producing a composite high-molecular polymer on various materials, such as noble metal particles, thereby regulating and controlling photo- and electro-activity of the materials.

Description

technical field [0001] The invention relates to a method for catalyzing polymerization reaction at a fixed point on a nanometer material based on DNA regulation and control of biological enzymes. The invention belongs to the field of nano polymer materials. Background technique [0002] High molecular polymers with π-electron backbones have excellent electrical conductivity and are widely used in residents' daily life. Traditional chemical synthesis methods are often carried out under polar pH conditions, resulting in low yields and many side reactions. At the same time, temperature control and pressure control are strict, and equipment investment is high, which is not conducive to expanding production. The biological enzyme is a kind of biocatalyst with catalytic function and adjustable activity. When catalyzing chemical reactions, biological enzymes have the advantages of high activity, strong specificity, adjustable activity, and mild reaction conditions. Therefore, t...

Claims

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Application Information

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IPC IPC(8): C12P13/00
CPCC12P13/00
Inventor 何丹农徐艳王萍金彩虹
Owner SHANGHAI NAT ENG RES CENT FORNANOTECH
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