Application method for diagnosing aptamer based on conventional gel electrophoresis

A nucleic acid aptamer and gel electrophoresis technology, which is applied in the field of biomedical detection and analysis, can solve the problems of special requirements for nucleic acid aptamers, expensive equipment, and no clinical diagnostic application of nucleic acid aptamers, and achieve practical value Large, simple and easy method, good application prospects

Inactive Publication Date: 2012-07-11
NANCHANG UNIV
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Problems solved by technology

[0006] At present, the methods for applying nucleic acid aptamers to target detection include flow cytometry, biochip technology, two-site binding assay (sandwich method), molecular beacons, biosensors, etc., but these methods may require expensive equipment, or There are special requirements for nucleic acid aptamers, or complex experimental techniques are required, and the practical value is poor, so far, there is no real clinical diagnostic application of nucleic acid aptamers

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  • Application method for diagnosing aptamer based on conventional gel electrophoresis
  • Application method for diagnosing aptamer based on conventional gel electrophoresis
  • Application method for diagnosing aptamer based on conventional gel electrophoresis

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Embodiment Construction

[0029] The method of the present invention will be further described below through a specific example of detection of liver cancer by nucleic acid aptamers in liver cancer serum based on neutral polyacrylamide gel electrophoresis.

[0030] 1. Materials.

[0031] 1. Nucleic acid aptamer: In the previous work of our research group, the nucleic acid aptamer AP-HCS-9-90 was screened out by SELEX technology with liver cancer mixed serum as the target, and was synthesized by Shanghai Sangon Company. The synthetic nucleic acid aptamer was mixed with binding buffer (20 mmol / L Hepes-Na, 120 mmol / L NaCl, 5 mmol / L KCl, 1 mmol / L CaCl 2 , 1 mmol / L MgCl 2 ) was dissolved, and prepared into a nucleic acid aptamer mother solution with a concentration of 10 pmol / μl, and stored at -20°C after aliquoting.

[0032] 2. Serum samples: collect peripheral blood samples (non-anticoagulant) from healthy people who come to the First Affiliated Hospital of Nanchang University with definite diagnosis ...

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Abstract

The invention relates to an application method for diagnosing an aptamer based on the conventional gel electrophoresis, and belongs to the technical field of the detection and analysis of biomedicine. The application method comprises the following steps of: incubating the aptamer and a blood serum sample, performing the conventional polyacrylamide gel electrophoresis and nucleic acid dyeing, observing a result under an ultraviolet lamp and collecting ultraviolet filtering images, measuring a gray-scale value of a dissociated aptamer strap by gel image analysis software, and analyzing difference between cases and contrast quantitatively to realize quantitative diagnosis. According to the application method, a liver cancer blood serum aptamer is used as a research model to be applied to liver cancer diagnosis, so the sensitivity, specificity and accuracy are up to over 90 percent. The application method is simple, convenient, easy, high in practical value and high in diagnostic efficiency, and has a bright application prospect.

Description

technical field [0001] The invention belongs to the technical field of biomedical detection and analysis. Background technique [0002] Nucleic acid aptamer (aptamer) is an artificial single-stranded oligonucleotide ligand of a biological target molecule, which is screened from a random single-stranded oligonucleotide library by SELEX (Systematic Evolution of Ligands by Exponential Enrichment) technology. The basic steps of nucleic acid aptamer screening are as follows: design and synthesize a random single-stranded oligonucleotide library; co-incubate the target with the library, so that the nucleic acid in the library that can specifically bind to the target forms a complex with the target, separates the latter and Elute the nucleic acid, use the eluted nucleic acid as a template for in vitro amplification, prepare a new random single-stranded oligonucleotide library, and start the next round of screening; repeat several rounds of screening (incubation, isolation, amplific...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
Inventor 张焜和徐国峰魏淑琴陈文学王婷曾琴吕农华张吉翔
Owner NANCHANG UNIV
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