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Application of Boronic Acid Derivatives Functionalized Fluorescent Probes in Detection of 5-Hydroxymethylcytosine

A technology of hydroxymethylcytosine and boronic acid derivatives is applied in the field of 2-phenyl-4-carboxyquinoline functionalized fluorescent composite sensing materials, which can solve the problem of increasing the DNA loss rate, difficult to avoid background factor interference, The operation process is cumbersome and other problems, to achieve the effect of simple method, high sensitivity, and overcoming the complexity of measuring instruments

Inactive Publication Date: 2018-07-10
SHAANXI NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, these fluorescent methods require at least two steps of purification, which increases the loss rate of DNA, and the operation process is cumbersome, and it is difficult to avoid interference from background factors.

Method used

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  • Application of Boronic Acid Derivatives Functionalized Fluorescent Probes in Detection of 5-Hydroxymethylcytosine
  • Application of Boronic Acid Derivatives Functionalized Fluorescent Probes in Detection of 5-Hydroxymethylcytosine
  • Application of Boronic Acid Derivatives Functionalized Fluorescent Probes in Detection of 5-Hydroxymethylcytosine

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Embodiment 1

[0027] The application of the boric acid derivative functionalized fluorescent probe in the detection of 5-hydroxymethylcytosine, the specific detection method consists of the following steps:

[0028] 1. Dissolve the standard sample of 5-hydroxymethylcytosine DNA (base sequence 5'-CTTAAGCCG(5hmC)AGGTACCTTCC-3', molecular weight 6372.2) in ultrapure water and prepare 100μmol / L 5-hydroxymethylcytosine Cytosine DNA in water. Add 1 μL 10000U / mL T4-β-glucosyltransferase solution into 32 μL 1.25×NEBuffer4, add 1 μL 2mmol / L uridine diphosphate glucose, 10 μL 100 μmol / L 5-hydroxymethylcytosine DNA aqueous solution, mix well, React for 1 hour at 37°C. After the reaction is over, return to normal temperature, add an equal volume of extract to the reaction solution (the extract is a mixture of phenol, chloroform, and isoamyl alcohol in a volume ratio of 25:24:1, pH 8.0), vigorously shake and mix for 15 minutes, centrifuge for 10 minutes, take the supernatant and repeat centrifugal wash...

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Abstract

The invention discloses an application of a boronic acid derivative functional fluorescent probe to 5-hydroxymethyl cytosine detection. The boronic acid derivative functional fluorescent probe is a polymethylacrylic acid epoxy propyl ester nano-ball with 2-(4-dyhydroxy borane) phenyl-4-quinoline supported on the surface. The fluorescent probe has the fluorescent characteristics of the 2-(4-dyhydroxy borane) phenyl-4-quinoline, retains good biocompatibility of the polymethylacrylic acid epoxy propyl ester nano-ball, can specifically adsorb glycosylated 5-hydroxymethyl cytosine and realizes simple, rapid, high-sensitivity and low-limit detection for the 5-hydroxymethyl cytosine.

Description

technical field [0001] The invention belongs to the technical field of detection of 5-hydroxymethylcytosine (5hmC) in DNA, in particular to a 2-(4-dihydroxyborane)phenyl-4-carboxyl group that can specifically recognize a cis-dihydroxy structure Quinoline (PBAQA) functionalized fluorescent composite sensing material (PBAQA-P GMA ) in the detection of 5hmC in DNA strands. Background technique [0002] 5-Methylcytosine (5mC) plays a crucial role in the regulation of gene expression, genomic imprinting, and X chromosome inactivation. It was recently found that 5mC is in the TET (ten eleven translocation) family dioxygenase Fe 2+ / α-ketoglutarate can be oxidized to 5hmC under the catalysis. 5hmC, as a new type of cytosine variant that was discovered in 1952 and rediscovered in 2009, has been widely studied, and has been recognized as the sixth base after 5mC. This base plays a vital role in biological and physiological processes. Studies have proved that 5hmC is an important...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C09K11/06G01N21/64
CPCC09K11/06C09K2211/1466G01N21/6486
Inventor 张静陈华艳潘炯秀张微魏静茹
Owner SHAANXI NORMAL UNIV