Line-1 gene methylation quantitative detection method

Abstract

The invention discloses a Line-1 gene methylation quantitative detection method, which comprises the steps of: (1) treating a to-be-detected sample to obtain a sample template; (2) designing and synthesizing Line-1 gene methylation primers and Line-1 gene unmethylated primers; (3) respectively using the methylation primers and the unmethylated primers in step (2) to perform a PCR amplification reaction on the sample template in step (1) so as to obtain corresponding PCR amplification products respectively; (4) treating the PCR amplification products in step (3) respectively to obtain corresponding standard templates; and (5) carrying out real-time fluorescence quantitative PCR detection on the sample template in step (1) and the standard templates in step (4) respectively, and calculating the degree of methylation. The method provided in the invention designs the methylation primers and unmethylated primers specific to the Line-1 gene so as to perform qualitative and quantitative detection on Line-1 gene methylation by means of a MSP technique and a real-time fluorescence quantitative PCR technique.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and in particular relates to a quantitative detection method for Line-1 gene methylation. Background technique [0002] In the proliferation and differentiation of tumor cells, gene mutation, disordered expression of DNA methylation, loss of LOH, etc. are the main reasons for the regulation disorder of oncogenes and tumor suppressor genes, and DNA methylation, as a very important epigenetic mechanism, is Participate in the above regulatory process and play a potential role in the formation of tumors. [0003] There are three main detection methods for gene abnormal DNA methylation: methylation-sensitive restriction endonuclease method, bisulfite sequencing method (BSP) and methylation-specific PCR method (MSP). [0004] Among them, the methylation-specific PCR method is currently the most widely used and preferred method for studying gene methylation. However, this method still has the foll...

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Problems solved by technology

However, this method still has the following disadvantages: (1) This method can only be used for qualitative research, that is, it can only determine whether there is methylation; if it is required to be quantitative. Other methods need to be used for further detection; (2) To distinguish the difference in the amount of amplification products of methylated primers and non-methylated primers, it is necessary to strictly control the conditions and cycle number of the PCR reaction

[0008] No method has been found to detect Line-1 gene methylation

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