DNA hydrogel based on biological mimic enzyme signal amplification and application thereof

A biosimulation and hydrogel technology, applied in the direction of chemical reaction analysis, chemical/physical process, physical/chemical process catalyst, etc., can solve food safety hazards and other problems, and achieve low cost and good portability , detect the effect of fast

Active Publication Date: 2020-06-19
INST OF ENVIRONMENTAL MEDICINE & OCCUPATIONAL MEDICINE ACAD OF MILITARY MEDICINE ACAD OF MILITARY SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In addition, MC-LR can also be enriched in aquatic products th

Method used

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  • DNA hydrogel based on biological mimic enzyme signal amplification and application thereof
  • DNA hydrogel based on biological mimic enzyme signal amplification and application thereof
  • DNA hydrogel based on biological mimic enzyme signal amplification and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0060] This example prepares a DNA hydrogel based on biomimetic enzyme signal amplification, in which the biomimetic enzyme is Cu / Au / Pt with peroxidase activity, and the outer hydrogel coating uses linear polymer The backbone of the material is a straight-chain polyacrylamide backbone. The following is the preparation method of the DNA hydrogel of this embodiment:

[0061] Step 1: Preparation method of Cu / Au / Pt TNPs:

[0062] Cu / Au / Pt TNPs biomimetic enzymes with peroxidase activity were synthesized by a "one-pot method". 12 μL CuSO 4 ﹒ 5H 2 O (0.1mol / L) and 25μL trisodium citrate (0.1mol / L) were added to 10mL ultrapure water, mixed well and then added 500μL KBH 4 (25mmol / L), after stirring for 15min, 25μL HAuCl 4 (0.1mol / L) and 25μL K 2 PtCl 4 (0.1mol / L) were added dropwise to the above mixture, and Cu / Au / Pt TNPs could be obtained after stirring for 20 min.

[0063] Step 2: Design of the first short-strand DNA and the second short-strand DNA

[0064] Design principl...

Embodiment 2

[0076] In this example, whether the binding competitiveness between the first short-strand DNA (SA) and the second short-strand DNA (SB) and the aptamer designed in Example 1 satisfies the points ①-② in the design principle verify. The verification method is as follows:

[0077] (1) Sample preparation:

[0078]

[0079] Note: Before adding MC-LR, denature the mixture at 95°C for 5 minutes, cool it to room temperature and place it at 4°C for 20 minutes. After adding MC-LR, incubate at 37°C for 40 minutes and store it in a -20°C refrigerator for testing.

[0080] (2) Take 200 μL of the samples in the above table respectively in a cuvette, and put them on a circular dichroism spectrometer for measurement. Circular dichroism instrument parameters are set as follows: measurement wavelength range: 200-350nm, scanning speed 100nm / min, response time 4s, bandwidth 2.0nm, scanning mode: continuous. Baseline correction: PBS solution.

[0081] (3) Data analysis:

[0082] Import th...

Embodiment 3

[0089] In this example, the DNA hydrogel based on the signal amplification of the biomimetic enzyme prepared in Example 1 is used for the quantitative detection of MC-LR in water, and the detection method is:

[0090] Take 10 μL of the Cu / Au / Pt-DNA hydrogel prepared in Example 1 into a 1.5 mL disposable centrifuge tube, add the sample solution to be tested (including MC-LR), and incubate at 37° C. for 40 min. Add 198 μL HO to the mixed system 2 O (pH=3), and the final concentrations of TMB and 48mmol / L of H 2 o 2 , After reacting at room temperature, concentrated hydrochloric acid was added to terminate the reaction. The absorbance value at a specific wavelength (optimally 452 nm) is measured by a UV-visible spectrophotometer.

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Abstract

The invention relates to DNA hydrogel based on biological mimic enzyme signal amplification and an application of the DNA hydrogel in microcystic toxin-LR detection. A DNA hydrogel coating layer is used for coating a biological mimic enzyme with peroxidase activity, and when the hydrogel coating layer is constructed, an aptamer of microcystin toxin-LR is used as a cross-linking bridge. Therefore,when the DNA hydrogel encounters the microcystic toxin-LR, the cross-linking bridge structure in the DNA hydrogel structure is changed, so that the DNA hydrogel is disintegrated to release the biological mimic enzyme, the biological mimic enzyme can catalyze a chromogenic reaction, and the concentration or content of the microcystic toxin-LR is detected by a colorimetric detection means. Comparedwith traditional detection methods such as high performance liquid chromatography (HPLC), enzyme-linked immunosorbent assay and liquid chromatography-tandem mass spectrometry, the method has the advantages of being low in cost, fast in detection, good in portability of matched detection equipment and the like.

Description

technical field [0001] The invention belongs to the technical field of water environment monitoring, and in particular relates to a DNA hydrogel based on biological simulation enzyme signal amplification and its application in microcystin-LR detection. Background technique [0002] Eutrophication of water bodies has led to more frequent cyanobacterial blooms in rivers and lakes around the world. The microcystins (MCs) produced and released by cyanobacteria such as Anabaena, Adenophora, Cylindrocystis and Nodularia have received extensive attention. MCs are a class of biologically active cyclic heptapeptide compounds that can exist stably in the environment, among which microcystin-LR (abbreviation: MC-LR) is the most abundant (accounting for about the total concentration of MCs in natural water bodies). 99.8%), the most toxic, has seriously threatened the ecological security of water bodies and human health. A large number of studies have shown that MC-LR exposure is highl...

Claims

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Application Information

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IPC IPC(8): B01J31/06G01N21/78
CPCB01J31/06G01N21/78Y02A50/30
Inventor 高志贤李双白家磊彭媛宁保安王江韩殿鹏任汉林韩铁伍翩
Owner INST OF ENVIRONMENTAL MEDICINE & OCCUPATIONAL MEDICINE ACAD OF MILITARY MEDICINE ACAD OF MILITARY SCI
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