High-throughput analysis of global dna methylation using methyl-sensitive digestion
- DNA methylation is a major regulatory process of gene transcription, and aberrant DNA methylation is associated with various diseases including cancer. Many compounds have been reported to modify DNA methylation states. Despite increasing interest in the clinical application of drugs with epigenetic effects, and the use of diagnostic markers for genome-wide hypomethylation in cancer, large-scale screening systems to measure the effects of drugs on DNA methylation are limited. In this study, we improved the previously established fluorescence polarization-based global DNA methylation assay so that it is more suitable for application to human genomic DNA. Our methyl-sensitive fluorescence polarization (MSFP) assay was highly repeatable (inter-assay coefficient of variation = 1.5%) and accurate (r2 = 0.99). According to signal linearity, only 50–80 ng human genomic DNA per reaction was necessary for the 384-well format. MSFP is a simple, rapid approach as all biochemical reactions and final detection can be performed in one well in a 384-well plate without purification steps in less than 3.5 hours. Furthermore, we demonstrated a significant correlation between MSFP and the LINE-1 pyrosequencing assay, a widely used global DNA methylation assay. MSFP can be applied for the pre-screening of compounds that influence global DNA methylation states and also for the diagnosis of certain types of cancer.
Author: | Hiromi Shiratori, Carmen Feinweber, Claudia Knothe, Jörn LötschORCiDGND, Dominique Jeanette ThomasORCiDGND, Gerd GeisslingerORCiDGND, Michael J. ParnhamORCiDGND, Eduard Resch |
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URN: | urn:nbn:de:hebis:30:3-417208 |
DOI: | https://doi.org/10.1371/journal.pone.0163184 |
ISSN: | 1932-6203 |
Pubmed Id: | https://pubmed.ncbi.nlm.nih.gov/27749902 |
Parent Title (English): | PLoS one |
Publisher: | PLoS |
Place of publication: | Lawrence, Kan. |
Contributor(s): | Sriharsa Pradhan |
Document Type: | Article |
Language: | English |
Date of Publication (online): | 2016/10/20 |
Date of first Publication: | 2016/10/17 |
Publishing Institution: | Universitätsbibliothek Johann Christian Senckenberg |
Release Date: | 2016/10/20 |
Volume: | 11 |
Issue: | (10): e0163184 |
Page Number: | 16 |
First Page: | 1 |
Last Page: | 16 |
Note: | Copyright: © 2016 Shiratori et al. This is an open access article distributed under the terms of the Creative Commons Attribution License http://creativecommons.org/licenses/by/4.0/, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. |
HeBIS-PPN: | 400651785 |
Institutes: | Medizin / Medizin |
Dewey Decimal Classification: | 5 Naturwissenschaften und Mathematik / 57 Biowissenschaften; Biologie / 570 Biowissenschaften; Biologie |
6 Technik, Medizin, angewandte Wissenschaften / 61 Medizin und Gesundheit / 610 Medizin und Gesundheit | |
Sammlungen: | Universitätspublikationen |
Licence (German): | Creative Commons - Namensnennung 4.0 |