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Aptamer of glycosylated hemoglobin and preparation method thereof

A technology of glycated hemoglobin and nucleic acid aptamer, which is applied in the field of protein detection, can solve the problems of lack of high-efficiency and specific recognition of glycated hemoglobin GHb, screening and preparation methods have not been reported, and achieve easy synthesis and labeling, which is beneficial to design and simplification The effect of the detection method

Active Publication Date: 2014-10-22
广西安仁欣生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, at present, there is still a lack of efficient and specific recognition research on glycosylated hemoglobin GHb based on nucleic acid aptamers, and the nucleic acid aptamers and their screening and preparation methods for glycosylated hemoglobin GHb have not been reported yet.

Method used

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  • Aptamer of glycosylated hemoglobin and preparation method thereof
  • Aptamer of glycosylated hemoglobin and preparation method thereof
  • Aptamer of glycosylated hemoglobin and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Example 1 Preparation of aptamer GHb1 specifically binding to glycated hemoglobin

[0040] (1) Construction of random sequence oligonucleotide library: artificially synthesized single-stranded DNA sequence, and the construction capacity of the library is 1×10 5 The single-stranded DNA random sequence oligonucleotide library, the DNA sequence included in the single-stranded DNA random oligonucleotide library is:

[0041] 5’-GACCATACCAGCTTATTCAATT-N 25~40 -AGATAGTAAGTGCAATCTGGC-3'

[0042] The DNA sequence of the single-stranded DNA random oligonucleotide library includes an intermediate random sequence N 25~60 and fixed sequences at both ends, the middle random sequence N 25~40 It is a random sequence of 25 to 40 bases, and the fixed sequences at both ends are: 5'-GACCATACCAGCTTATTCAATT, 3'-CGGTCTAACGTGAATGATAGA, and the fixed sequences at both ends are PCR amplification primer binding regions;

[0043] (2) Preparation of glycosylated hemoglobin-micro-magnetic...

Embodiment 2

[0049] Example 2 Preparation of aptamer GHb2 specifically binding to glycosylated hemoglobin

[0050] (1) Construction of random sequence oligonucleotide library: artificially synthesized single-stranded DNA sequence, and the construction capacity of the library is 3×10 5 The single-stranded DNA random sequence oligonucleotide library, the DNA sequence included in the single-stranded DNA random oligonucleotide library is:

[0051] 5’-GACCATACCAGCTTATTCAATT-N 30~45 -AGATAGTAAGTGCAATCTGGC-3'

[0052] The DNA sequence of the single-stranded DNA random oligonucleotide library includes an intermediate random sequence N 25~60 and fixed sequences at both ends, the middle random sequence N 30~45 It is a random sequence of 30 to 45 bases, and the fixed sequences at both ends are: 5'-GACCATACCAGCTTATTCAATT, 3'-CGGTCTAACGTGAATGATAGA, and the fixed sequences at both ends are PCR amplification primer binding regions;

[0053] (2) Preparation of glycosylated hemoglobin-micro-magne...

Embodiment 3

[0059] Example 3 Preparation of aptamer GHb3 specifically binding to glycosylated hemoglobin

[0060] (1) Construction of random sequence oligonucleotide library: artificially synthesized single-stranded DNA sequence, and the construction capacity of the library is 1×10 6 The single-stranded DNA random sequence oligonucleotide library, the DNA sequence included in the single-stranded DNA random oligonucleotide library is:

[0061] 5’-GACCATACCAGCTTATTCAATT-N 35~60 -AGATAGTAAGTGCAATCTGGC-3'

[0062] The DNA sequence of the single-stranded DNA random oligonucleotide library includes an intermediate random sequence N 25~60 and fixed sequences at both ends, the middle random sequence N 35~60 It is a random sequence of 35 to 60 bases, the fixed sequences at both ends are: 5'-GACCATACCAGCTTATTCAATT, 3'-CGGTCTAACGTGAATGATAGA, and the fixed sequences at both ends are PCR amplification primer binding regions;

[0063] (2) Preparation of glycosylated hemoglobin-micro-magnetic...

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Abstract

The invention discloses an aptamer with high specificity and affinity of glycosylated hemoglobin GHb and a preparation method of the aptamer. The nucleotide sequence of the aptamer of the glycosylated hemoglobin GHb comprises any one of sequences from SEQ ID NO: 1 to SEQ ID NO: 4. The preparation method comprises the following steps of: constructing a random sequence oligonucleotide library, preparing a glycosylated hemoglobin-micro magnetic bead composite, initially screening a target material-nucleic acid composite, preparing a secondary single stranded DNA oligonucleotide library, screening and identifying the aptamer of the glycosylated hemoglobin. The prepared aptamer is free from toxicity, small in molecular weight, and liable to synthesize and mark. The aptamer only specifically identifies the glycosylated hemoglobin GHb, and does not identify other homologous non-glycosylated hemoglobin Hb, so that the aptamer can be taken as a molecular beacon for detecting the content of the glycosylated hemoglobin GHb.

Description

[0001] This application is a divisional application of "a nucleic acid aptamer for glycosylated hemoglobin and its preparation method". Nucleic acid aptamer and its preparation method". technical field [0002] The invention belongs to the technical field of protein detection and relates to a nucleic acid, in particular to a nucleic acid aptamer of glycosylated hemoglobin GHb with high specificity and high affinity and a preparation method thereof. Background technique [0003] Diabetes is a worldwide public health problem of human health. Diabetes is a lifelong disease, and its complications are the main cause of disability and death. Therefore, people hope that diabetes can be detected and treated as early as possible. Traditional diabetes diagnosis and treatment monitoring use fasting blood glucose, postprandial blood glucose and oral glucose tolerance test, etc., but blood glucose parameters only represent the instantaneous blood glucose level when blood is drawn, and gl...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/115C12N15/10G01N33/72
Inventor 李寿东胡建红李戈强李朝君谢志峰李朝志
Owner 广西安仁欣生物科技有限公司
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