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Method for detecting 5-aldehyde cytosine and 5-hydroxymethylcytosine in DNA (deoxyribonucleic acid) by utilizing piperidine aqueous solution

A technology of hydroxymethylcytosine and aldehydecytosine, which is applied in the field of DNA sequencing, can solve the problem of rarely being detected

Inactive Publication Date: 2013-04-24
WUHAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

A lot of articles on DNA methylation detection have been reported, but 5fC, which was just discovered in 2011, has rarely been detected

Method used

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  • Method for detecting 5-aldehyde cytosine and 5-hydroxymethylcytosine in DNA (deoxyribonucleic acid) by utilizing piperidine aqueous solution
  • Method for detecting 5-aldehyde cytosine and 5-hydroxymethylcytosine in DNA (deoxyribonucleic acid) by utilizing piperidine aqueous solution

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0014] Example 1: Reaction of 5-formylcytosine DNA with 5% piperidine

[0015] Take 2 μL of 5-formylcytosine DNA (10 μM), add 93 μL of water and 5 μL of piperidine, heat-treat at 90°C for 0.5 hours, spin the solvent, load the sample to the electrophoresis tank, and observe the gel results after 2 hours , 10% of the 5-formylcytosine DNA was fragmented.

Embodiment 2

[0016] Example 2: Reaction of 5-formylcytosine DNA with 10% piperidine

[0017] Take 2 μL of 5-formylcytosine DNA (10 μM), add 88 μL of water and 10 μL of piperidine, heat-treat at 90°C for 0.5 hours, spin the solvent, load the sample to the electrophoresis tank, and observe the gel results after 2 hours , 35% of the 5-formyl cytosine DNA fragmentation occurred.

Embodiment 3

[0018] Example 3: Reaction of 5-formylcytosine DNA with 15% piperidine

[0019] Take 2 μL of 5-formylcytosine DNA (10 μM), add 83 μL of water and 15 μL of piperidine, heat-treat at 90°C for 0.5 hours, spin the solvent, load the sample to the electrophoresis tank, and observe the gel results after 2 hours , 50% of the 5-formylcytosine DNA fragmentation occurred.

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Abstract

The invention discloses a method for detecting 5-aldehyde cytosine and 5-hydroxymethylcytosine in DNA (deoxyribonucleic acid) by utilizing a piperidine aqueous solution. Piperidine specifically reacts with 5-aldehyde cytosine in DNA with a fluorescent mark; the 5-aldehyde cytosine can be detected while the 5-aldehyde cytosine in the DNA is oxidized into 5-aldehyde cytosine through potassium perruthenate, and subjected to piperidine treatment, and the position and the quantity of the 5-hydroxymethylcytosine can be obtained by comparing two reacting DNA bands in a DNA polyacrylamide modified adhesive. By adopting the method, detection of the 5-aldehyde cytosine and the 5-hydroxymethylcytosine can be rapidly, effectively and sensitively achieved.

Description

technical field [0001] The invention relates to a method for detecting 5-formylcytosine and 5-hydroxymethylcytosine in DNA by using a piperidine aqueous solution, belonging to the field of DNA sequencing. Background technique [0002] As an important branch of genetics, epigenetics mainly studies changes in gene expression levels based on non-gene sequence changes, and its main contents include DNA methylation, genomic imprinting, RNA interference, and histone modification. In recent years, more and more people have begun to pay attention to DNA methylation, and related research has been carried out in full swing. DNA methylation is one of the earliest discovered modification pathways, and its existence is closely related to gene silencing and expression. Usually, under the action of methyltransferase, methylation occurs at the C5 position of cytosine to form 5-methylcytosine (5mC), and then after treatment with TET family enzymes, 5mC is converted into 5-hydroxymethylc...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 周翔毛伍祥郭枫晚
Owner WUHAN UNIV
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