A kind of ruthenium complex and its preparation method and its application in detecting 5-formylcytosine

A formylcytosine and ruthenium complex technology, applied in the field of analysis and detection, can solve the problems of DNA sample degradation, limited resolution, low expression level, etc., and achieve the effect of simple conditions, wide response range, and fewer synthesis steps

Active Publication Date: 2021-06-15
SUN YAT SEN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Antibody-based DNA immunoprecipitation and high-throughput sequencing (DIP-seq) is a simple and efficient method for the analysis of cytosine modifications (especially for detecting regions with many modified bases), but this method requires Highly specific antibodies, there are still some difficulties in recruiting highly specific antibodies, and the detection resolution is limited
Single-base resolution technology based on bisulfite sequencing, which requires bisulfite treatment, can easily lead to degradation of DNA samples, and requires deeper sequencing depths to reliably detect 5fC
Given the low level of expression of 5fC in the genome, it will be challenging to map 5fC at a genome-wide scale and at fundamental resolution

Method used

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  • A kind of ruthenium complex and its preparation method and its application in detecting 5-formylcytosine
  • A kind of ruthenium complex and its preparation method and its application in detecting 5-formylcytosine
  • A kind of ruthenium complex and its preparation method and its application in detecting 5-formylcytosine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] The synthesis of embodiment 1 ruthenium complex Ru-1C

[0051] (1) Weigh bis-(2,2'-bipyridine) (1,10-o-phenanthroline-5,6-dione) ruthenium (II) (1equ) hexafluorophosphate and dissolve it in acetonitrile (MeCN ), drop a few drops of acetic acid; dissolve 3,4-diaminobenzyl alcohol in acetonitrile and drop it into the previous system, cool after reflux for 2 hours, add excess saturated potassium hexafluorophosphate solution, centrifuge to obtain an orange-red precipitate, After drying in vacuum, purify by recrystallization to obtain bis-(2,2'-bipyridyl) hexafluorophosphate (bipyridyl[3,2-a:2',3'-c]phenazin-11-ylmethanol) Ruthenium(II);

[0052] (2) Di-(2,2'-bipyridyl) (dipyridyl[3,2-a:2',3'-c]phenazin-11-ylmethanol) obtained after the above purification ) ruthenium(II) was dissolved in acetonitrile, and excess phosphorus tribromide was added, after stirring at room temperature for 4 hours, an excess saturated potassium hexafluorophosphate solution was added, and an orang...

Embodiment 2

[0058] The synthesis of embodiment 2 ruthenium complexes Ru-2C

[0059] (1) Weigh bis-(2,2'-bipyridyl)(1,10-o-phenanthrene-5,6-dione)ruthenium(II)(1equ) hexafluorophosphate and dissolve it in DMF , drop a few drops of acetic acid; dissolve 2-(3,4-diaminophenoxy)ethyl bromide (1.5equ) in DMF first and then drop it into the previous system, reflux for 2 hours and cool, then add excess saturated six Potassium fluorophosphate solution, centrifuged to obtain an orange-red precipitate, vacuum dried, recrystallized and purified to obtain bis-(2,2'-bipyridyl)(11-(2-bromoethoxy)-bipyridyl[3, 2-a: 2',3'-c] phenazine) ruthenium (II);

[0060] (2) Then weigh bis-(2,2'-bipyridyl)(11-(2-bromoethoxy)-bipyridyl[3,2-a:2',3'-c]phenone hexafluorophosphate Azine) ruthenium (II), dissolved in DMF, adding excess sodium carbonate, then adding N-hydroxyphthalimide, stirring at room temperature overnight, adding excess saturated potassium hexafluorophosphate solution, and centrifuging to obtain an o...

Embodiment 3

[0063] The synthesis of embodiment 3 ruthenium complexes Ru-4C

[0064](1) Weigh bis-(2,2'-bipyridyl)(1,10-o-phenanthrene-5,6-dione)ruthenium(II)(1equ) hexafluorophosphate and dissolve it in DMF , drop a few drops of acetic acid; dissolve 2-(3,4-diaminophenoxy)butyl bromide (1.5equ) in DMF first and drop it into the previous system, reflux for 2 hours and cool, add excess saturated six Potassium fluorophosphate solution, centrifuged to obtain an orange-red precipitate, after vacuum drying, recrystallized and purified to obtain bis-(2,2'-bipyridine)(11-(2-bromobutoxy)-bipyridine[3, 2-a: 2',3'-c] phenazine) ruthenium (II);

[0065] (2) Then weigh bis-(2,2'-bipyridyl)(11-(2-bromobutoxy)-bipyridyl[3,2-a:2',3'-c]phenone hexafluorophosphate Azine) ruthenium (II), dissolved in DMF, adding excess sodium carbonate, then adding N-hydroxyphthalimide, stirring at room temperature overnight, adding excess saturated potassium hexafluorophosphate solution, and centrifuging to obtain an ora...

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Abstract

The invention discloses a novel ruthenium complex, its preparation method and its application in detecting 5-formylcytosine. The invention provides a novel ruthenium complex and a preparation method thereof. The invention also provides a fluorescent molecular probe for detecting 5-formylcytosine. The present invention also provides a method for detecting 5-formylcytosine by using the novel ruthenium complex or the ruthenium complex prepared by the preparation method or the fluorescent molecular probe. The ruthenium complex can uniquely combine with 5fC in DNA, thereby specifically recognizing 5fC in double-stranded DNA, has good fluorescence response and chemical stability, and has good thermodynamic stability, and can realize 5fC in double-stranded DNA. Real-time detection and intracellular distribution and positioning, low cost, low equipment requirements, no need to enrich 5-formylcytosine, direct detection, and wide application conditions, the conditions are simpler, milder, and high sensitivity.

Description

technical field [0001] The invention belongs to the technical field of analysis and detection. More specifically, it relates to a ruthenium complex and its preparation method and its application in the detection of 5-formylcytosine. It designs and synthesizes a class of polypyridine ruthenium complexes, and uses the characteristic functional groups of the ligands on the complexes to interact with 5- Formylcytosine forms a bond, enabling specific detection of 5-formylcytosine in double-stranded DNA. Background technique [0002] DNA is an important carrier of genetic information in living organisms. It contains four nitrogenous bases, adenine (A), guanine (G), cytosine (C) and thymine (T). In addition to the four natural bases, bases with modified groups have also been found. These modification sites are located at the 2nd carbon of adenine, the 5th carbon of cytosine and the methyl group of the 5th carbon of thymine. base, and these modification groups are located in the m...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07F15/00C09K11/06G01N21/64
CPCC07F15/0053C09K11/06C09K2211/185G01N21/6486
Inventor 徐亮赖培冬林娇
Owner SUN YAT SEN UNIV
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