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E-cadherin methylation detection method based on hydrosulphite sequencing method

A technology of bisulfite and cadherin, which is applied in the direction of biochemical equipment and methods, microbial measurement/inspection, etc., can solve the problems of high failure rate, improve the success rate, improve specificity and sensitivity, and reduce non-specific The effect of heterosexual amplification

Inactive Publication Date: 2016-08-17
廖静
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Problems solved by technology

[0003] The present invention provides a method for detecting cadherin methylation based on bisulfite sequencing to solve the problem of high failure rate in the prior art

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  • E-cadherin methylation detection method based on hydrosulphite sequencing method
  • E-cadherin methylation detection method based on hydrosulphite sequencing method
  • E-cadherin methylation detection method based on hydrosulphite sequencing method

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Embodiment Construction

[0025] Embodiments of the invention are described in detail below, but the invention can be practiced in many different ways as defined and covered by the claims.

[0026] Since DNA methylation is closely related to the occurrence and development of tumors, the detection of abnormal DNA methylation sites of specific genes is one of the hot spots in the research of tumor biomarkers. After the genomic DNA to be tested is converted by bisulfite, unmethylated cytosine is converted to uracil, while 5-methylated cytosine remains unchanged. That is to say, the vast majority of methylation detection methods convert the subtle changes of chemical molecules before and after modification into base changes and display them visually through PCR amplification. However, excess genomic DNA is likely to cause incomplete bisulfite modification, resulting in the inability of partially unmethylated cytosine to be effectively converted into uracil, resulting in false positive results.

[0027] Af...

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Abstract

The invention discloses an E-cadherin methylation detection method based on a hydrosulphite sequencing method. The E-cadherin methylation detection method includes the steps that firstly, rats which are 6-7 weeks old are selected, the kidneys of the killed rats are collected, connective tissue is sheared off, the renal cortex is taken and sheared into 50-100 mg tissue fragments, and the tissue fragments are quick-frozen with liquid nitrogen and stored in a -80 DEG C refrigerator; secondly, genome DNA is extracted with a genome DNA extraction reagent kit; thirdly, the concentration and purity of genome DNA are measured with a nucleic acid quantometer, the integrity of genome DNA is detected through 0.5%-1.0% sepharose gel electrophoresis, and DNA samples detected to be qualified are placed in a -20 DEG C--80 DEG C refrigerator for use; fourthly, genome DNA is modified with a methylation conversion reagent kit, and the concentration of modified genome DNA is determined with the nucleic acid quantometer; fifthly, PCR amplification is carried out under different experiment conditions; sixthly, samples used in follow-up steps are selected in the fifth step. By means of the E-cadherin methylation detection method, the success rate of E-cadherin methylation experiments is increased.

Description

technical field [0001] The invention relates to the biological field, in particular to a method for detecting cadherin methylation based on a bisulfite sequencing method. Background technique [0002] Currently, bisulfite sequencing is the gold standard for the detection of deoxyribonucleic acid (DNA) methylation sites recognized by scholars at home and abroad, which can accurately and sensitively detect specific DNA sequences with different cytosine guanine dinucleotide (CpG ) site methylation status, but the low efficiency of polymerase chain reaction (PCR) amplification that frequently occurs during the experiment is one of the important reasons for the stagnation of the experiment. Contents of the invention [0003] The invention provides a method for detecting cadherin methylation based on a bisulfite sequencing method to solve the problem of high failure rate in the prior art. [0004] In order to solve the above problems, as an aspect of the present invention, a me...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
CPCC12Q1/6869C12Q1/6848C12Q2531/113C12Q2523/125C12Q2527/137
Inventor 廖静李小峰
Owner 廖静
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