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Updated: Aug 3, 2026
Enhanced Reduced Representation Bisulfite Sequencing for Assessment of DNA Methylation at Base Pair Resolution
Published on: February 24, 2015
A capture methyl-seq protocol with improved efficiency and cost-effectiveness using pre-pooling and enzymatic
Keita Hasegawa1,2, Kazuhiko Nakabayashi3, Keisuke Ishiwata1
1Department of Maternal-Fetal Biology, National Center for Child Health and Development, 2-10-1 Okura, Setagaya-ku, Tokyo, 157-8535, Japan.
Objective:
The opportunities for sequencing-based methylome analysis of clinical samples are increasing. To reduce its cost and the amount of genomic DNA required for library preparation, we aimed to establish a capture methyl-seq protocol, which adopts pre-pooling of multiple libraries before hybridization capture and TET2/APOBEC-mediated conversion of unmethylated cytosine to thymine.
Results:
We compared a publicly available dataset generated by the standard Agilent protocol of SureSelect XT Human Methyl-Seq Kit and our dataset obtained by our modified protocol, EMCap, that adopted sample pre-pooling and enzymatic conversion. We confirmed that the quality of DNA methylation data was comparable between the two datasets. As our protocol, EMCap, is more cost-effective and reduces the amount of input genomic DNA, it would serve as a better choice for clinical methylome sequencing.
