Method for establishing real-time fluorescent quantitative detection of PER2 promoter methylation level

Abstract

The invention discloses a method for establishing a real-time fluorescent quantitative detection PER2 promoter methylation level. The method comprises the following steps of S1, selecting a PER2 promoter CpG island region: 1) querying a PER2 promoter sequence according to a gene database, and 2) predicting a GpG island enrichment region contained in the PER2 promoter sequence through a CpG island online prediction website, and determining a target sequence, S2, designing a primer and a probe for PER2 promoter methylation detection according to the target sequence, S3, designing methylation and non-methylation sequences of a target sequence according to a transformation rule of bisulfite on a gene sequence, artificially synthesizing corresponding sequences, and storing the corresponding sequences in a plasmid form, and S4, establishing a real-time fluorescent quantitative PCR detection PER2 promoter methylation method, and confirming the performance of the PER2 promoter methylation method. The methylation level of the PER2 promoter can be accurately and quantitatively detected, and an effective tool and method are provided for researching PER2 methylation.

Description

technical field [0001] The invention relates to the technical field of detecting the methylation level of the PER2 promoter, in particular to a method for establishing real-time fluorescent quantitative detection of the methylation level of the PER2 promoter. Background technique [0002] Many studies have proved that hypermethylation of PER2 gene occurs in leukemia patients, especially myeloid leukemia patients, and PER2 methylation may be an important epigenetic marker for early diagnosis, prognosis and treatment of leukemia. At present, the methylation-specific MSP method is mostly used in the detection of PER2 promoter methylation, which is a qualitative detection method and is not easy to be used for dynamic monitoring and level comparison. At present, there is no method for quantitative detection of PER2 gene promoter methylation that has undergone strict performance verification, and there is no reference material that can be used for quantitative detection of PER2 pr...

AI Technical Summary

Problems solved by technology

However, the sulfite pyrosequencing method is expensive

[0004] F Hernandez-Rosas (PMID: 30008892) et al. used methylation-specific PCR (MS-PCR) to detect the methylation of the PER2 promoter, but this method can only be used for qualitative detection and cannot be accurately quantified or dynamically detected. changes in the level of

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