Ratio electrochemical DNA sensor-modified electrode for detecting gene P53 and preparation method of modified electrode

A technology for modifying electrodes and DNA strands, applied in the field of electrochemical detection, can solve the problems of high equipment investment, limited clinical application, complicated operation, etc., and achieve the effects of low cost of use, convenient operation, and simple reaction conditions

Active Publication Date: 2017-02-22
QINGDAO UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] Existing detection methods for the P53 gene mainly include capillary electrophoresis, denaturing high-performance liquid chromatography, denaturing gradient gel electrophoresis, and allele function analysis techniques separated from yeast. Although these methods have high sensitivity, they are complicated to operate. , high cost and many other disadvantages; at the same time, the high investment cost of instruments and expensive consumables also limit the clinical application of these methods

Method used

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  • Ratio electrochemical DNA sensor-modified electrode for detecting gene P53 and preparation method of modified electrode
  • Ratio electrochemical DNA sensor-modified electrode for detecting gene P53 and preparation method of modified electrode
  • Ratio electrochemical DNA sensor-modified electrode for detecting gene P53 and preparation method of modified electrode

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Effect test

Embodiment 1

[0041] A preparation method for the modified electrode of the P53 gene based on the double hairpin structure, such as figure 1 shown, including the following steps:

[0042] (1) Design a single-stranded DNA auxiliary probe S1, the bases at both ends of the DNA chain can be complementary to the bases at both ends of the capture probe chain S2 and have a complementary paired sequence of 10 bases in length, which can be fixed on A hairpin structure is formed behind the surface of the gold electrode; the 5' end of the S1 chain is modified with a ferrocene label, which can generate electrochemical signals; the 53' end of the S1 chain is thiolated so that S1 can be modified by a gold-sulfur bond to the surface of the gold electrode; the DNA strand sequence of the auxiliary probe S1 is: 5'-Fc-CTCTCA GTG ATT TTT TTA GTG AGA GAG-(CH 2 ) 6 -SH-3';

[0043] (2) Design a single-stranded DNA capture probe S2, the bases in the middle part of the DNA chain can be complementary paired with...

Embodiment 2

[0049] A preparation method for the modified electrode of the P53 gene based on the double hairpin structure, such as figure 1 shown, including the following steps:

[0050] (1) Design a single-stranded DNA auxiliary probe S1, the bases at both ends of the DNA chain can be complementary to the bases at both ends of the capture probe chain S2 and have a complementary paired sequence of 10 bases in length, which can be fixed on A hairpin structure is formed on the surface of the gold electrode; the 5' end of the S1 chain is modified with a ferrocene label, which can generate electrochemical signals; the 53' end of the S1 chain is thiolated, so that S1 is modified to the Gold electrode surface; wherein, the DNA strand sequence of the auxiliary probe S1 is: 5'-Fc-CTCTCA GTG ATT TTT TTA GTG AGA GAG-(CH 2 ) 6 -SH-3';

[0051] (2) Design a single-stranded DNA capture probe S2, the bases in the middle part of the DNA chain can be complementary paired with the P53 gene; at the same ...

Embodiment 3

[0057] The application of the modified electrode prepared in Example 1 in detecting the P53 gene, the steps are as follows:

[0058] Soak the modified electrode of Example 1 in Tris-HCl buffer solution, the Tris-HCl buffer solution is a mixture containing 110-150mM Tris-HCl, 100-120mM NaCl, 1-10mM MgCl, its pH is 7.4, water bath Heat for 15 minutes. Then form a three-electrode system with a calomel reference electrode and a platinum wire contrast electrode, and perform DPV detection in Tris-HCl buffer solution to obtain the peak current I of methylene blue 0(MB) , the peak current of ferrocene is I 0(Fc) , repeated several times to take the average value.

[0059] Configure a series of p53 gene buffer solutions, in which the concentration and pH value of the buffer solution are kept constant, and the concentration of p53 gene is adjusted to 10 -8 , 5×10 -9 , 10 -9 , 5×10 -10 , 10 -10 , 5×10 -11 , 10 -11 , 10 -12 M, then soak the obtained electrode in the buffer solut...

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Abstract

The invention relates to the technical field of electrochemical detection, and particularly discloses a modified electrode for detecting a tumor suppressor gene P53 and a preparation method of the modified electrode. A probe for detecting the gene P53 is composed of a single-stranded DNA auxiliary probe S1 and a single-stranded DNA capturing probe S2, wherein the DNA strand sequence of the S1 is 5'-Fc-CTC TCA GTG ATT TTT TTA GTG AGA GAG-(CH2)6-SH-3', and the DNA strand sequence of the S2 is 5'-MB-TCA CTG AGT CTT CCA GTG TGA TGA TCA CT-3'. According to the method, preparation is easy, control is convenient, the use cost is low, and compared with the methods such as capillary electrophoresis, denaturing high performance liquid chromatography, denaturing gel gradient electrophoresis and a yeast-separated allele function analysis technique, the electrochemical biosensor mode has the advantages that samples do not need to be pretreated, control is easy, the reaction conditions are simple, and the cost is low.

Description

technical field [0001] The invention relates to the technical field of electrochemical detection, in particular to a ratio electrochemical DNA sensor modified electrode for detecting tumor suppressor gene P53 and a preparation method thereof. Background technique [0002] The tumor suppressor gene P53 is a tumor suppressor gene known as "the guardian of the genome", and it is a homologous gene of a series of isoform proteins (P53 protein). A series of proteins encoded by this gene can regulate cell division, repair and apoptosis. When a cell is damaged, the P53 protein monitors the degree of damage to the cell's genes. When the gene damage is small, the P53 protein will prompt the cell to repair itself; and when the damaged gene cannot be repaired, the P53 protein will induce cell apoptosis. Therefore, when the P53 gene is mutated, the process of cell repair and apoptosis will be disturbed, so that some damaged cells will continue to divide and grow under abnormal conditio...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11G01N27/26G01N27/327
CPCC12Q1/6825G01N27/26G01N27/327C12Q2565/519C12Q2565/607
Inventor 夏建飞徐姣艳王宗花龚世达孙华
Owner QINGDAO UNIV
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