DNA methylation is one of the most widely studied changes in the field of epigenetics. This biochemical phenomenon exerts a pervasive hand in a multitude of biological processes, including deactivation of chromosome X, genomic imprinting, stem cell differentiation, gene expression control and chromosome stability. DNA methylation, with its labyrinthine reach, assumes a pivotal role in orchestrating the symphony of gene expression and presiding over an array of biological processes and pathologies. At present, there are many methods to determine the methylation of DNA samples. In this article, we will explore the techniques used for DNA methylation analysis, with a focus on bisulfite treatment-based methods.
Fig 1. The main method for 5-methyl cytosine analysis after treatment with bisulfite. (Khodadadi E, et al.,2021)
Sequence-Based Analysis
In the contemporary landscape of scientific inquiry, the advent of high-throughput sequencing technologies has ushered in a transformative era for the analysis of DNA methylation patterns. Within this paradigm, two prominent methodologies have emerged as stalwart contenders: whole-genome bisulfite sequencing (WGBS) and reduced representation bisulfite sequencing (RRBS).
WGBS provides a comprehensive view of the entire methylome, allowing researchers to identify methylation patterns across the entire genome. RRBS, on the other hand, offers a cost-effective alternative by selectively sequencing regions of interest, such as promoter regions or CpG islands.
Analysis Based on Melting Temperature
Melting temperature analysis is grounded in a fundamental principle: the dissimilarities in melting temperatures exhibited by methylated and unmethylated DNA strands. This analytical approach involves the systematic application of ascending temperatures to DNA samples, yielding distinctive melting curves that differentiate between the methylated and unmethylated variants.
This method is particularly useful for rapid screening of DNA methylation patterns and can be employed in clinical settings for diagnostic purposes. Nevertheless, it is vital to acknowledge that this method may, at times, fall short in terms of discerning the intricate nuances discernible through sequence-based analysis, rendering it better suited for preliminary assessments.
Interaction-Based Analysis
In the domain of epigenetics, specialized analytical techniques delve into the intricate interplay between DNA methylation and other epigenetic modifications, such as histone modifications. Chromatin immunoprecipitation (ChIP) followed by bisulfite sequencing (ChIP-BS) is one such method that combines DNA methylation analysis with the mapping of histone modifications. This synergy provides profound insights into the coordinated orchestration of gene expression through epigenetic modifications.
Additionally, interaction-based analysis can uncover the role of DNA methylation in mediating protein-DNA interactions. For instance, methylated CpG dinucleotides can recruit specific proteins, leading to altered gene expression patterns. Techniques like methylated DNA immunoprecipitation (MeDIP) followed by sequencing (MeDIP-seq) are designed to identify such interactions and their functional implications.
In conclusion, these techniques collectively contribute to our understanding of DNA methylation and its role in health and disease. As technology continues to advance, we can expect even more powerful and precise methods for unraveling the secrets of DNA methylation.
Reference
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Khodadadi E, Fahmideh L, Khodadadi E, et al. Current Advances in DNA Methylation Analysis Methods. Biomed Res Int. 2021;2021:8827516.
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