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Aptamer of sfrp1 protein and its screening method and application

A protein and sequence technology, applied in the field of SFRP1 protein aptamer and its screening and application, can solve the problem of lack of seven transmembrane domains

Active Publication Date: 2017-03-15
灏灵赛奥(天津)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

[0002] SFRP, secreted frizzled-related proteins (secret frizzled-related proteins), contains a cysteine-rich domain (cysteine ​​rich domain, CRD), but lacks seven transmembrane domains, it may be related to frizzled, Frz) competes with Wnt protein, thereby inhibiting the overexpression of Wnt protein

Method used

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  • Aptamer of sfrp1 protein and its screening method and application
  • Aptamer of sfrp1 protein and its screening method and application
  • Aptamer of sfrp1 protein and its screening method and application

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Experimental program
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Embodiment 1

[0013] 1. Preparation of SFRP1 protein

[0014] The SFRP1 protein was obtained by means of yeast recombinant expression well known to those skilled in the art, and the protein sequence is shown in SEQ ID NO: 1; the concentration of the protein solution was 15 mg / ml.

[0015] 2. Synthesis of libraries and primers

[0016] 2.1. Synthesize ssDNA oligonucleotide library for screening (5′-TCA GTC GCT TCG CCG TCT CCTTC----N35----GCA CAA GAG GGA GAC CCC AGA GGG-3′), wherein N35 is 35 a random oligonucleotide;

[0017] Primer P1: TCAGTCGCTTCGCCGTCTCCTTC;

[0018] Primer P2: CCCTCTGGGGTCTCCCTCTTGTGC.

[0019] 2.2. SELEX screening of aptamers, the specific method is as follows:

[0020] 2.2.1 The binding and separation of ssDNA and SFRP1 protein, the specific method is as follows:

[0021] Take 4 μL of 100 μM ssDNA oligonucleotide library, dilute to 100 μl with 2× binding buffer, denature at 95°C for 5 minutes, add 100 μl SFRP1 protein after 10 minutes in ice bath, combine on a sha...

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Abstract

The invention relates to a group of oligonucleotide sequences capable of recognizing SFRP1 protein and preparation methods thereof. The oligonucleotide sequences comprise SEQ ID No. 2-16, have high affinity and specificity and can be used for detection of SFRP1 protein.

Description

Background technique [0001] Oligonucleotide aptamers are screened by SELEX technology (Systematic evolution of ligands by exponential enrichment) biological library technology. The principle of this technology is to use molecular biology techniques to construct artificially synthesized single-stranded random oligonucleotide libraries. The length of its random sequence is about 20-100 bases. Utilizing the flexible and changeable molecular conformation of single-stranded oligonucleotides, the random oligonucleotide library interacts with target molecules, retains oligonucleotides that bind to target molecules in a spatial conformation, and undergoes repeated amplification and screening After a few cycles, the oligonucleotide sequences that specifically bind to the target can be enriched, and finally the specific oligonucleotide aptamers, ie, aptamers, of various target molecules can be obtained. The pattern of aptamer recognition molecules screened by SELEX technology is simila...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/435C12N15/115C12Q1/68G01N33/68
CPCC07K14/47C12N15/115C12N2310/16G01N33/6863
Inventor 荆东辉
Owner 灏灵赛奥(天津)生物科技有限公司