Method for detecting transcription factor by biosensor and glucometer

A biosensor and transcription factor technology, applied in the biological field, can solve problems such as difficult to meet quantitative needs, difficult to popularize the application of ordinary people, expensive instruments, etc., and achieve the effects of rapid and convenient detection, wide application range and high sensitivity

Inactive Publication Date: 2013-09-04
GUANGZHOU INST OF BIOMEDICINE & HEALTH CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although compared with traditional methods, these new technologies are easy to operate and have high detection sensitivity, but they all require laboratory-specific instruments and professional technicians, and the instruments are expensive, making it difficult to promote them to the general population.
Test strip biosensors have also been reported for the detection of transcription factors, but they can only provide qualitative analysis, which is difficult to meet the actual quantitative needs, thus limiting its practical application (Z.Fang, C.Ge, W.Zhang, P. Lie and L. Zeng, Biosens. Bioelectron., 2011, 27, 192-196)

Method used

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  • Method for detecting transcription factor by biosensor and glucometer
  • Method for detecting transcription factor by biosensor and glucometer
  • Method for detecting transcription factor by biosensor and glucometer

Examples

Experimental program
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Effect test

Embodiment 1

[0024] Embodiment 1: Utilize the method described in the present invention to detect transcription factor Oct4

[0025] The detection principle of this embodiment is as follows figure 1 shown. Biotin-modified double-stranded DNA (double-stranded DNA, dsDNA) was immobilized on streptavidin (SA)-modified magnetic beads (magneticbeads, MBs) (through the interaction of SA-biotin), The excess DNA was removed by magnetic bead separation or washed with water to obtain the SA-MBs-biotindsDNA complex. The sequence of the double-stranded DNA is shown in Table 1. The double-stranded DNA sequence contains the binding site of the transcription factor Oct4 (the underlined part in the double-stranded DNA sequence in Table 1).

[0026] Table 1

[0027] name

dna sequence

biotin-modified DNA

5'-biotin-CATTGTT ATGCAAAT CAGTC-3'

SEQ ID NO:1

Complementary DNA

3'-GTAACAA TACGTTTA GTCAG-5'

SEQ ID NO:2

[0028] Then add transcription ...

Embodiment 2

[0035]Embodiment 2: Using the method of the present invention to detect transcription factor Oct4

[0036] The detection principle of this embodiment is as follows figure 1 shown. Immobilize biotin-modified double-stranded DNA (double-stranded DNA, dsDNA) (through the interaction of SA-biotin) on streptavidin (streptavidin, SA)-modified microtiter plate or electrode), and water Wash to remove excess DNA to obtain SA-microplate / electrode-biotin dsDNA complex. The sequence of the double-stranded DNA is shown in Table 1 (see Example 1).

[0037] Then add transcription factor Oct4 to SA-microplate / electrode-biotin dsDNA, it will bind to double-stranded DNA to form SA-microplate / electrode-biotin dsDNA-Oct4 complex, wash with water to remove excess Oct4 , then add anti-Oct4 antibody (anti-Oct4antibody, Ab) and invertase (invertase) modified gold nanoparticles (gold nanoparticles, AuNPs), then the Ab / invertase-AuNPs complex is immobilized on the enzyme label through antibody-Oct4 ...

Embodiment 3

[0041] Example 3: Detection of transcription factor Oct4 at different concentrations using the method of the present invention

[0042] Prepare standard solutions of transcription factor Oct4 at concentrations of 0.05ng / mL, 0.1ng / mL, 0.5ng / mL, 2.5ng / mL, 12.5ng / mL and 25ng / mL, and store at room temperature.

[0043] Add different concentrations of Oct4 solutions to the reaction system described in Example 1, react fully, and finally take 5 μL of the supernatant for detection with a blood glucose meter, and record the reading of the blood glucose meter after 25 seconds.

[0044] From figure 2 It can be seen from the detection results that when Oct4 of 0.05ng / mL exists, the corresponding reading can be read on the blood glucose meter, indicating that the detection limit of Oct4 by the biosensor of the blood glucose meter is 0.05ng / mL, which is higher than that of the traditional ELISA method. The sensitivity is more than 20 times higher. Moreover, as the concentration of Oct4 ...

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Abstract

The invention relates to a method for detecting a transcription factor by a biosensor and a glucometer. The method comprises the following steps of 1, fixing a nucleic acid sequence specifically binding to a transcription factor on a solid matrix, and adding a sample into the solid matrix with the nucleic acid sequence to obtain a compound, 2, fixing a transcription factor antibody and invertase on carriers, 3, mixing the solid matrix obtained by the step 1 and the carriers obtained by the step 2, and removing the unbound carriers, and 4, adding excess cane sugar into the mixture obtained by the step 3, carrying out a reaction for a certain time, detecting glucose content by a conventional glucometer, and converting the glucose content into content of the transcription factor in the sample. The method has high sensitivity and singularity, simple processes, a low cost and a fast detection rate, adopts the portable detection apparatuses, and is suitable for popular popularization.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to a method for detecting a transcription factor, in particular to a method for detecting a transcription factor by using a biosensor and a blood glucose meter. Background technique [0002] Transcription factors are a class of DNA-binding proteins that can bind to specific DNA sequences and play an important role in the regulation of gene expression. They are closely related to tumorigenesis, gene repair, cell differentiation and apoptosis. Traditional methods for the detection of DNA-binding proteins mainly include: gel migration retention assay (electrophoretic mobility shift assay, EMSA), enzyme-linked immunosorbent assay (enzyme-linked immunosorbent assay, ELISA), DNA footprinting assay (DNA footprinting) and Western blotting. However, these methods are cumbersome to operate, time-consuming and labor-intensive, and are not conducive to the rapid detection of transcription factors. ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/53
Inventor 曾令文陈俊华吴薇
Owner GUANGZHOU INST OF BIOMEDICINE & HEALTH CHINESE ACAD OF SCI
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