Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Method for detecting methylation level of synuclein intron 1

A synuclein and level detection technology, applied in the field of bioengineering, can solve the problems of decreased quantitative accuracy, time-consuming and laborious, and incapable of quantitative analysis, and achieve the effects of good quantitative characteristics, convenient operation, and high throughput

Active Publication Date: 2013-09-25
CHANGCHUN HENGXIAO BIOTECH CO LTD
View PDF0 Cites 4 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Among them, the sulfite sequencing method involves a large number of amplified fragment cloning work, which is time-consuming and laborious.
Also, the ratio of methylated / unmethylated products may shift during the cloning process, resulting in a loss of quantitative accuracy
The methylation-specific PCR method is relatively simple, but this method can only be used qualitatively, not quantitatively
The next-generation sequencing method is expensive and not suitable for large-scale applications

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
  • Method for detecting methylation level of synuclein intron 1
  • Method for detecting methylation level of synuclein intron 1
  • Method for detecting methylation level of synuclein intron 1

Examples

Experimental program
Comparison scheme
Effect test

Embodiment

[0041] Example: Application of Pyrosequencing in α-Syn Intron 1 Methylation Detection

[0042] 1. Sodium bisulfite treatment

[0043] Genomic DNA is modified using sodium metabisulfite and hydroquinone to convert unmethylated cytosines to uracils, while methylated uracils remain unchanged. The process is briefly described as follows: first, the genomic DNA was denatured with 0.3mol / L NaOH; then, the genomic DNA was treated at 50°C for 16h with a mixed solution of sodium metabisulfite and hydroquinone (pH 5.0). The DNA was purified with a kit, and 0.3 mol / L NaOH was added to incubate at 37°C for 15 min to terminate the reaction, and finally the DNA was precipitated with ethanol.

[0044] 2. PCR amplification

[0045] Perform PCR amplification according to conventional methods, primers and reaction conditions are as follows:

[0046] Forward amplification primers: GAA, ATG, GAA, GTG, TAA, GGA, GGT, T;

[0047] Reverse amplification primers: Biotin-TTC, TAA, TCC, ATC, CAA, CA...

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 relates to a method for detecting the methylation level of a synuclein intron 1 and belongs to the field of bioengineering. The method comprises the following detection steps of: performing sodium hydrogen sulfite treatment; performing polymerase chain reaction (PCR) amplification; preparing a single chain; preparing a pyrophosphoric acid sequencing sample; preparing before pyrophosphoric acid sequencing analysis; analyzing and calculating the methylation state of a corresponding locus by using computer software. The deoxyribonucleic acid (DNA) methylation detection method provided by the invention overcomes the defects in the existing DNA methylation detection technology and has the advantages of quickness, accuracy, high flux and quantitative property. A method for detecting the CpG island methylation level of an alpha-Syn intron 1 based on a pyrophosphoric acid sequencing platform is established. Electrophoresis, Sanger sequencing and pyrophosphoric acid sequencing prove the detection specificity of the method disclosed by the invention. Compared with a sulfite clone sequencing method, the pyrophosphoric acid sequencing method disclosed by the invention is proved to have a high quantitative property. In addition, the method is convenient and quick to operate and high in flux. 96 samples can be sequenced simultaneously, and the time consumption is less than 2 hours, so the method is extremely suitable for clinical examination work.

Description

technical field [0001] The invention belongs to the field of biological engineering. Background technique [0002] Currently, sulfite sequencing, methylation-specific PCR, and next-generation sequencing are mainly used to detect the methylation level of α-Syn intron 1. Among them, the sulfite sequencing method involves a large amount of amplified fragment cloning work, which is time-consuming and labor-intensive. Also, the ratio of methylated / unmethylated products may shift during cloning, resulting in a loss of quantitative accuracy. The methylation-specific PCR method is relatively simple, but this method can only be used qualitatively, not quantitatively. The next-generation sequencing method is expensive and not suitable for large-scale applications. Contents of the invention [0003] The purpose of the present invention is to provide a detection method capable of quantitatively analyzing the methylation level of CpG island in α-Syn intron 1. [0004] Detection ste...

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
Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
Inventor 蔡延宁于顺关恒
Owner CHANGCHUN HENGXIAO BIOTECH CO LTD
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

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

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com