Identifying DNA methylation in a nanochannel

Sakon Rahong,Noritada Kaji,Tomoji Kawai,Takao Yasui,Yoshinobu Baba,Kazuki Nagashima,Masaki Kanai,Xiaoyin Sun,Takeshi Yanagida

doi:10.1080/14686996.2016.1223516

Sakon Rahong, Noritada Kaji + Show 7 more

Open Access

https://doi.org/10.1080/14686996.2016.1223516

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Abstract

DNA methylation is a stable epigenetic modification, which is well known to be involved in gene expression regulation. In general, however, analyzing DNA methylation requires rather time consuming processes (24–96 h) via DNA replication and protein modification. Here we demonstrate a methodology to analyze DNA methylation at a single DNA molecule level without any protein modifications by measuring the contracted length and relaxation time of DNA within a nanochannel. Our methodology is based on the fact that methylation makes DNA molecules stiffer, resulting in a longer contracted length and a longer relaxation time (a slower contraction rate). The present methodology offers a promising way to identify DNA methylation without any protein modification at a single DNA molecule level within 2 h.

Highlights

DNA methylation is a stable epigenetic modification that it is known to be involved in the regulation of gene expression
DNA methylation detection in previous studies is based on modification of methylation sites using methyl-CpG-binding domain (MBD) proteins, which has a risk of non-specific absorption to DNA molecules
The nanochannel device was fabricated on fused silica substrates by photolithography, reactive ion etching and electron beam lithography techniques

Summary

Introduction

DNA methylation is a stable epigenetic modification that it is known to be involved in the regulation of gene expression. We fabricated a nanochannel device to detect the DNA methylation without site specificity at a single DNA molecule level by measuring a contraction process (Figure 1). To measure the contraction process, we introduced methylated and non-methylated single DNA molecules into the nanochannel to elongate and contract them (Figure 2(a)). After elongation of DNA molecules, we turned off the electrical potential to evaluate the contraction process of DNA molecules with and without methylation inside the nanochannel. This simple analysis method helps to detect DNA molecules with methylation

Device fabrication

Sample preparation

Measurement setup

Results and discussion

Conclusions