Specific marking method for 5-hydroxymethyluracil on DNA (deoxyribonucleic acid)

A technology of hydroxymethyluracil and phosphorothioatemethyluracil, which is applied in the field of specific labeling of 5-hydroxymethyluracil on DNA, can solve the problem of large demand for detection samples, harsh chemical reaction conditions, and low reaction efficiency. Low-level problems, to achieve the effect of improving accuracy and sensitivity, easy to obtain reaction reagents, and simple reaction conditions

Active Publication Date: 2020-05-01
XI AN JIAOTONG UNIV
View PDF2 Cites 5 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] In order to overcome the shortcomings of the above-mentioned prior art, the object of the present invention is to provide a method for specific labeling of 5-hydroxymethyl

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Specific marking method for 5-hydroxymethyluracil on DNA (deoxyribonucleic acid)
  • Specific marking method for 5-hydroxymethyluracil on DNA (deoxyribonucleic acid)
  • Specific marking method for 5-hydroxymethyluracil on DNA (deoxyribonucleic acid)

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Example 1: Using the specific labeling method for 5-hmU of the present invention to realize the distinction of different bases.

[0044] Prepare dsDNA oligonucleotide sequences containing different bases (including restriction endonuclease NcoI site 5'-CCAXGG-3', X=T, U, hmU and fU), and catalyze the primer synthesis by DNA polymerase in vitro For extension, dsDNA model substrates containing sites of different base pairs (5hmU:A, 5hmC:G, 5fC:G, 5fU:A, U:A or T:A) were prepared.

[0045] Wherein, in order to form 5hmU:G-containing dsDNA, the 5-hmU:A-containing dsDNA product was incubated with a G template to perform a strand displacement reaction. Product characterization in model DNA labeling reactions by 5-HMUDK and SH reaction reagents. Such as figure 2 Shown is the melting curve of the DNA sample cleaved by the restriction endonuclease NcoI, and it can be seen that the reaction product 5psmU can block the cleavage of DNA by NcoI.

[0046] 5-HMUDK thiophosphorylat...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention discloses a specific marking method for 5-hydroxymethyluracil on DNA (deoxyribonucleic acid) and belongs to the technical field of molecular biology. The specific marking method comprises the following steps: transferring gamma-thiophosphate to 5-hmU from 5-O-thiotriphosadenine by using 5-hmU separated from bacteriophage M6 of pseudomonas aeruginosa so as to generate 5-thiophosphoricacid methyluracil, representing the generated 5-thiophosphoric acid methyluracil by using a liquid chromatogram-tandem mass spectrometry method, and marking sulfydryl through crosslinking chemistry,so as to achieve specific marking on 5-hydroxymethyluracil on DNA. By adopting the method disclosed by the invention, defects that a great amount of detection samples are needed, chemical reaction conditions are harsh, the reaction efficiency is low, and the like, can be effectively overcome.

Description

technical field [0001] The invention belongs to the technical field of molecular biology and relates to a specific labeling method of 5-hydroxymethyluracil on DNA. Background technique [0002] In addition to the four classic bases, the genome also contains chemically modified DNA bases. These modified bases can be produced by endogenous enzymes or exogenous factors, and they have the potential to profoundly affect genome function and cellular processes. Many modified DNA bases have been discovered, among which 5-methylcytosine (5mC) and its oxidized derivative 5-hydroxymethylcytosine (5-hydroxymethylcytosine, 5mC) are the most famous in mammalian genomes. -hmC), 5-fluorocytosine (5-formylcytosine, 5-fC) and 5-carboxycytosine (5-carboxylcytosine, 5-caC). These epigenetic marks have been shown to play important roles in regulating gene expression. Since the methylation modification of DNA was discovered in 1948, it has always been a research hotspot in the field of epigene...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
IPC IPC(8): C12N15/11C12Q1/6844
CPCC12N15/11C12N2310/10C12Q1/6844C12N2310/315C12N2330/30C12Q2531/119
Inventor 赵永席陈锋白敏张进
Owner XI AN JIAOTONG UNIV
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap