Elsevier

Analytical Biochemistry

Volume 317, Issue 2, 15 June 2003, Pages 259-265
Analytical Biochemistry

Polymerase chain reaction-based methods of DNA methylation analysis

https://doi.org/10.1016/S0003-2697(03)00169-6Get rights and content

Abstract

DNA methylation is the main epigenetic modification in humans, and changes in methylation patterns play an important role in tumorigenesis. Hypermethylation of normally unmethylated CpG islands in the promoter regions often occurs in important tumor suppressor genes, DNA repair genes, and metastasis inhibitor genes. The changes of methylation status of various gene promoters seem to be a common feature of malignant cells and these changes can occur early in the progression process. Therefore detection of aberrant promoter hypermethylation of cancer-related genes may be useful for cancer diagnosis or detection of cancer recurrence. The purpose of this review is to provide a summary of the most commonly used techniques for the study of DNA methylation. Current scientific literature involving methylation detection methods was reviewed with an emphasis on polymerase chain reaction (PCR)-based detection methods. The current methodologies may be broadly classed into PCR-based methylation assays and non-PCR-based methylation assays. The problems and advantages of the different methods for detecting aberrant methylation are discussed. As the number of genes known to be hypermethylated in cancer is growing, the detection of aberrant promoter region methylation will be a promising approach for using DNA-based markers for the early detection of human cancers. Many techniques, especially PCR-based methylation assay techniques, make it practical to use these new methylation biomarkers in early cancer diagnosis.

Section snippets

PCR-based methods of DNA methylation analysis

Cytosines located 5 to guanines, called CpG dinucleotides, are present in the regulatory regions of many genes. In normal cells, CpG dinucleotides in gene transcriptional promoter regions are generally unmethylated. However, in the promotor sequences of genes associated with certain cancers or inherited diseases these cytosines are methylated. The methylation status of some of these promoter regions can provide important information with respect to the nature of a tissue or cellular specimen.

Critical note to these analytical approaches with respect to clinical application

PCR-based sensitive detection methods made it possible to assess methylation changes in cancer cells. These PCR-based strategies to assess methylation changes in minute quantities of biological materials raise the possibility that this type of analysis could facilitate risk assessment or early diagnosis strategies [29]. This strategy is further enhanced by three facts: (1) the PCR signal is a positive signal that cannot be masked through contamination by normal cells, (2) promoter

Conclusions

As the number of genes known to be hypermethylated in cancer is growing, the detection of aberrant promoter region methylation will be a promising approach for using DNA-based markers for the early detection of human cancers. Many techniques to make methylation detection possible have been developed over the years [34], [35]. New methods and approaches to avoid certain limitations or to be more practical are developed continually [36], [37], [38], [39], [40]. Therefore, so many techniques

Acknowledgements

This study was supported in part by the Japan–China Sasakawa Medical Fellowship.

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