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Preparation method and application of electrochemical sensor for detecting transgenic soybeans based on double-signal output of gene editing technology

A technology of genetically modified soybeans and gene editing, which is applied in the fields of biochemical equipment and methods, electrochemical variables of materials, and analysis by making materials undergo chemical reactions. High, selective, and easy-to-operate effects

Active Publication Date: 2021-07-09
NINGBO UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, there are no relevant reports on the detection of genetically modified soybeans by electrochemical biosensors based on the CRISPR / Cas12 system at home and abroad, and there are no relevant reports on the detection of genetically modified products with dual signal output of ECL and FSCV.

Method used

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  • Preparation method and application of electrochemical sensor for detecting transgenic soybeans based on double-signal output of gene editing technology
  • Preparation method and application of electrochemical sensor for detecting transgenic soybeans based on double-signal output of gene editing technology
  • Preparation method and application of electrochemical sensor for detecting transgenic soybeans based on double-signal output of gene editing technology

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Experimental program
Comparison scheme
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Embodiment 1

[0038] A method for preparing an electrochemical sensor for detecting transgenic soybeans based on gene editing technology with dual signal output, such as figure 1 shown, including the following steps:

[0039] 1. Magnetic material Fe 3 o 4 Synthesis of @AuNPs

[0040] (1) Add 2.8 g FeCl 3 ·6H 2 O, 7.8 g NH 4 Ac and 0.8 g of sodium citrate were dissolved in 135 mL of ethylene glycol, stirred vigorously at 155 °C for 1 h under nitrogen protection, then transferred to a Teflon-lined stainless steel autoclave, heated at 200 °C for 15 h, and then naturally Cool to room temperature to obtain magnetic beads, wash the magnetic beads with ethanol and ultrapure water several times, and disperse them in 50 mL of ethanol to obtain ferric oxide nanoparticles Fe 3 o 4 NPs;

[0041] (2) Add 45 mL Fe 3 o 4 NPs were mixed with 0.5 mL 3-aminopropyltriethoxysilane (APTES), sonicated for 0.5 hours, stirred at room temperature for 7 hours, magnetically separated, washed, and redispers...

Embodiment 2

[0051]With above-mentioned embodiment 1, its difference is:

[0052] Step 1 Magnetic Material Fe 3 o 4 @AuNPs is synthesizing:

[0053] (1) Add 2.5g FeCl 3 ·6H 2 O, 7.5 g NH 4 Ac and 0.5 g of sodium citrate were dissolved in 150 mL of ethylene glycol, stirred vigorously at 140 °C for 1 h under nitrogen protection, then transferred to a Teflon-lined stainless steel autoclave, heated at 200 °C for 14 h, and then Naturally cool to room temperature to obtain magnetic beads, wash the magnetic beads with ethanol and ultrapure water several times, and disperse them in 50 mL of ethanol to obtain ferric oxide nanoparticles Fe 3 o 4 NPs;

[0054] (2) Add 40 mL Fe 3 o 4 NPs were mixed with 0.4 mL of 3-aminopropyltriethoxysilane (APTES), sonicated for 0.5 hours, stirred at room temperature for 5 hours, magnetically separated, washed, and redispersed in 50 mL of water to obtain aminated Fe 3 o 4 NPs;

[0055] (3) Add 0.6 mL of 0.3 mol / L NaOH solution and 0.5 mL of 80 wt% tetrak...

Embodiment 3

[0061] With above-mentioned embodiment 1, its difference is:

[0062] Step 1 Magnetic Material Fe 3 o 4 @AuNPs is synthesizing:

[0063] (1) Add 3.0 g FeCl 3 ·6H 2 O, 8.0 g NH 4 Ac and 1.0 g of sodium citrate were dissolved in 150 mL of ethylene glycol, stirred vigorously at 170 °C for 1 h under nitrogen protection, then transferred to a Teflon-lined stainless steel autoclave, heated at 200 °C for 16 h, and then Naturally cool to room temperature to obtain magnetic beads, wash the magnetic beads with ethanol and ultrapure water several times, and disperse them in 50 mL of ethanol to obtain ferric oxide nanoparticles Fe 3 o 4 NPs;

[0064] (2) Add 50 mL Fe 3 o 4 NPs were mixed with 0.6 mL 3-aminopropyltriethoxysilane (APTES), sonicated for 0.5 hours, stirred at room temperature for 8 hours, magnetically separated, washed, and redispersed in 50 mL of water to obtain aminated Fe 3 o 4 NPs;

[0065] (3) Add 1.0 mL of 0.1 mol / L NaOH solution and 0.7 mL of 80 wt% tetrahy...

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Abstract

The invention discloses a preparation method and application of an electrochemical sensor for detecting transgenic soybeans based on double-signal output of a gene editing technology. The preparation method is characterized by comprising the following steps: synthesizing a magnetic material Fe3O4@AuNPs; a step of synthesizing the signal unit Fe3O4@AuNPs&DNA-Fc / [Ru(bpy)2]<2+; uniformly mixing a Cas12a enzyme solution and a crRNA solution, incubating at room temperature, adding a t-DNA solution and a binding buffer solution, uniformly mixing, incubating at room temperature, adding a signal unit solution, sucking a magnetic material part by adopting a magnetic separation method after cutting is finished, and re-dispersing the magnetic material part into 20 [mu]L of water; and finally, dripping the magnetic material part on the surface of the treated magnetic glassy carbon electrode, and obtaining the electrochemical sensor, detecting transgenic soybeans through the rapid scanning voltammetry technology and the electrochemical luminescence method, and therefore, the electrochemical sensor has the advantages of being high in sensitivity and selectivity and easy and rapid to operate.

Description

technical field [0001] The present invention relates to a method for detecting transgenic soybeans, in particular to a preparation method and application of an electrochemical sensor for detecting transgenic soybeans based on dual-signal output of gene editing technology. Background technique [0002] As an emerging biotechnology, transgenic technology refers to the use of genetic engineering to add desired target genes to the genetic material of other organisms, thereby improving the original traits of the organisms or endowing them with new excellent traits. In recent years, the application and development of transgenic technology in the field of agriculture has been very rapid, and it has become the focus of competition in agricultural science and technology around the world. At present, transgenic soybean is an important food crop in the world, accounting for 57% (58.6 million hectares) of the total planting area of ​​transgenic crops in the world. At the same time, the...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N27/327G01N27/30G01N27/48G01N21/76C12Q1/6804
CPCG01N27/3278G01N27/308G01N27/48G01N21/76C12Q1/6804C12Q2521/327C12Q2525/161
Inventor 葛浩然郭智勇汪小福徐俊锋郝婷婷陈小双
Owner NINGBO UNIV
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