Whole-genome Methylation Analysis of APOBEC Enzyme-converted DNA (~5 kb) by Nanopore Sequencing
Suzuko Zaha,
Yoshitaka Sakamoto,
Satoi Nagasawa,
Sumio Sugano,
Ayako Suzuki,
Yutaka Suzuki,
Masahide Seki
Affiliations
Suzuko Zaha
Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Kashiwa, Chiba, Japan
Yoshitaka Sakamoto
Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Kashiwa, Chiba, Japan
Satoi Nagasawa
Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Kashiwa, Chiba, Japan
Sumio Sugano
Institute of Kashiwa-no-ha Omics Gate, Kashiwa, Chiba, JapanFuture Medicine Education and Research Organization, Chiba University, Chuo, Chiba-city, Chiba, Japan
Ayako Suzuki
Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Kashiwa, Chiba, Japan
Yutaka Suzuki
Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Kashiwa, Chiba, Japan
Masahide Seki
Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Kashiwa, Chiba, Japan
In recent years, DNA methylation research has been accelerated by the advent of nanopore sequencers. However, read length has been limited by the constraints of base conversion using the bisulfite method, making analysis of chromatin content difficult. The read length of the previous method combining bisulfite conversion and long-read sequencing was ~1.5 kb, even using targeted PCR. In this study, we have improved read length (~5 kb), by converting unmethylated cytosines to uracils with APOBEC enzymes, to reduce DNA fragmentation. The converted DNA was then sequenced using a PromethION nanopore sequencer. We have also developed a new analysis pipeline that accounts for base conversions, which are not present in conventional nanopore sequencing, as well as errors produced by nanopore sequencing.
