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Updated: Mar 26, 2026

DNA Methylation: Bisulphite Modification and Analysis
Published on: October 21, 2011
A simple modification to improve the accuracy of methylation-sensitive restriction enzyme quantitative polymerase
Magdalena Krygier1, Justyna Podolak-Popinigis2, Janusz Limon1
1Department of Biology and Genetics, Medical University of Gdańsk, 80-210 Gdańsk, Poland.
Abstract:
DNA digestion with endonucleases sensitive to CpG methylation such as HpaII followed by polymerase chain reaction (PCR) quantitation is commonly used in molecular studies as a simple and inexpensive solution for assessment of region-specific DNA methylation. We observed that the results of such analyses were highly overestimated if mock-digested samples were applied as the reference. We determined DNA methylation levels in several promoter regions in two setups implementing different references: mock-digested and treated with a restriction enzyme that has no recognition sites within examined amplicons. Fragmentation of reference templates allowed removing the overestimation effect, thereby improving measurement accuracy.
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