Optical means for detecting i-motif conformation of DNA

An optical method, conformational technology, applied in biochemical equipment and methods, fluorescence/phosphorescence, microbial determination/inspection, etc., can solve the problem of long time required, achieve high selectivity and sensitivity, shorten detection time, The effect of simplifying the detection steps

Active Publication Date: 2009-05-20
SHANGHAI INST OF APPLIED PHYSICS - CHINESE ACAD OF SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, these detection methods require large-scale instruments and take a long time

Method used

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  • Optical means for detecting i-motif conformation of DNA
  • Optical means for detecting i-motif conformation of DNA
  • Optical means for detecting i-motif conformation of DNA

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Take 2.5 μL of C-probe solution with a concentration of 100 μM and add it to 2.5 μL of solutions with pH values ​​of 5.0, 6.8, 7.0 and 8.5, react at room temperature for 10 minutes, and then add PMNT solution of equal charge (1 mM PMNT solution 5 μL), observe the color change of the solution, and then measure the ultraviolet-visible absorption spectrum. At the same time, take any sequence of nc-probe solutions for negative control experiments.

[0038] The results showed that when the specific C-probe was used for detection, the color in the pH 5.0 solution was yellow, while in the other three solutions, the color was orange-red. This is due to the fact that residue C of DNA is protonated to form a semi-protonated C-C in a weakly acidic environment at pH 5.0 + base pair, thus forming a stable i-motif structure, which turns yellow when electrostatically combined with positively charged PMNT. In a neutral alkaline environment with a pH greater than or equal to 6.8, the ...

Embodiment 2

[0041] Take 2.5 μL of C-probe solution with a concentration of 100 μM and add it to 2.5 μL of the solution whose pH was adjusted with HCl and NaOH. Then add an isoelectric PMNT solution (5 μL of 1 mM PMNT solution) to it, and observe the color change of the solution, so as to investigate the influence on the detection of structural transformation in a strong acid or strong alkali environment.

[0042] The results showed that in the environment with pH less than 6.5 such as pH 1, 3 and 5, the color of the solution was yellow due to the formation of i-motif structure, while in the environment with pH greater than 6.5 such as pH 7, 9, 11 and 13 , the color of the solution is red because the i-motif structure cannot be formed. It shows that the detection of conformational changes of C-rich DNA can also be carried out in more acidic or alkaline environments.

Embodiment 3

[0044] Experiments were carried out according to the conditions in Example 1, and the reversibility of the method of the present invention for conformational transition detection was analyzed. First, the specific probe C-probe solution (sample 1) was added to the pH 8.0 solution. Since the four-chain structure cannot be formed, the color is red after adding the conjugated polymer. Then add a certain amount of strong acid solution (1M HCl) therein to make the system change to pH 5.0, the system is weakly acidic (sample 2), the DNA forms a four-stranded i-motfi structure, and the color turns yellow. After the color is stable, add 1M NaOH to make the solution pH become 8.0 (sample 3) again, and the color turns red again, then according to the above method, the solution pH becomes 5.0 (sample 4) again, repeat the experiment, and finally detect each The UV-Vis absorption spectrum of the solution.

[0045] see results image 3 . When the pH of the system is acidic, the color of ...

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Abstract

The invention discloses an optical method for detecting the four-chain-motif conformation of DNA, comprising the following steps: 1) the cationic water-soluble conjugated polymer solution with equivalent charge amount is added to DNA solution; 2) and then colorimetric and / or fluorescence detection are / is carried out to the DNA solution, and finally the C-rich sequence conformation of the DNA to be detected, such as human telomere DNA, can be judged according to the color change of the solution, fluorescence spectra or UV-visible absorption spectrum of the solution. The invention has simple method and high selectivity and sensitivity, and has important significance in biological detection, early diagnosis and treatment of diseases.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to an optical method for detecting the conformation of the four-strand i-motif of DNA. Background technique [0002] The structure of the DNA molecule folds differently under different conditions. The diversity of DNA molecular structure plays a key role in human understanding and understanding of various life phenomena. Therefore, studying the structural transformation of DNA is of great significance in biology. [0003] Oligodeoxynucleotides rich in cytosine (cytosine, C) can form a stable non-B-DNA structure called "i-motif" structure under acidic conditions with pH ≤ 6.5. This structure is similar to but different from B-DNA. It is a tetramer formed by equivalent four strands, with a semi-protonated C-C inserted between two antiparallel double strands. + base pairs, thus binding together in a zipper, with the distance between the nearest base pairs being only 3.1 , than B-DNA (...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68G01N21/25G01N21/64
Inventor 刘兴奋王丽华樊春海
Owner SHANGHAI INST OF APPLIED PHYSICS - CHINESE ACAD OF SCI
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