Application of 5-Methylcytosine DNA Glycosylase to the Quantitative Analysis of DNA Methylation
1
Department of Agriculture, Forestry and Bioresources, Seoul National University, Seoul 08826, Korea
2
Interdisciplinary Program in Agricultural Genomics, Seoul National University, Seoul 08826, Korea
3
Research Institute of Agriculture and Life Sciences, Seoul National University, Seoul 08826, Korea
4
Plant Genomics and Breeding Institute, Seoul National University, Seoul 08826, Korea
*
Author to whom correspondence should be addressed.
†
Present address: Center for Genome Engineering, Institute for Basic Science, Daejeon 34126, Korea.
Int. J. Mol. Sci. 2021, 22(3), 1072; https://doi.org/10.3390/ijms22031072
Received: 25 December 2020 / Revised: 14 January 2021 / Accepted: 20 January 2021 / Published: 22 January 2021
(This article belongs to the Section Molecular Biology)
In higher eukaryotes DNA methylation is a prominent epigenetic mark important for chromatin structure and gene expression. Thus, profiling DNA methylation is important for predicting gene expressions associated with specific traits or diseases. DNA methylation is achieved by DNA methyltransferases and can be actively removed by specific enzymes in a replication-independent manner. DEMETER (DME) is a bifunctional 5-methylcytosine (5mC) DNA glycosylase responsible for active DNA demethylation that excises 5mC from DNA and cleaves a sugar-phosphate bond generating a single strand break (SSB). In this study, DME was used to analyze DNA methylation levels at specific epialleles accompanied with gain or loss of DNA methylation. DME treatment on genomic DNA generates SSBs in a nonsequence-specific fashion proportional to 5mC density, and thus DNA methylation levels can be easily measured when combined with the quantitative PCR (qPCR) method. The DME-qPCR analysis was applied to measure DNA methylation levels at the FWA gene in late-flowering Arabidopsis mutants and the CNR gene during fruit ripening in tomato. Differentially methylated epialleles were successfully distinguished corresponding to their expression levels and phenotypes. DME-qPCR is proven a simple yet effective method for quantitative DNA methylation analysis, providing advantages over current techniques based on methylation-sensitive restriction digestion.
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Keywords:
DNA methylation; DEMETER; DNA demethylase; epiallele; epigenetic profiling
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MDPI and ACS Style
Choi, W.L.; Mok, Y.G.; Huh, J.H. Application of 5-Methylcytosine DNA Glycosylase to the Quantitative Analysis of DNA Methylation. Int. J. Mol. Sci. 2021, 22, 1072. https://doi.org/10.3390/ijms22031072
AMA Style
Choi WL, Mok YG, Huh JH. Application of 5-Methylcytosine DNA Glycosylase to the Quantitative Analysis of DNA Methylation. International Journal of Molecular Sciences. 2021; 22(3):1072. https://doi.org/10.3390/ijms22031072
Chicago/Turabian StyleChoi, Woo L.; Mok, Young G.; Huh, Jin H. 2021. "Application of 5-Methylcytosine DNA Glycosylase to the Quantitative Analysis of DNA Methylation" Int. J. Mol. Sci. 22, no. 3: 1072. https://doi.org/10.3390/ijms22031072
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