Single-molecule real-time sequencing (SMRT) is used in the epigenomic and epitranscriptomic fields. As regards epigenomics, thousands of zero-mode waveguides (ZMWs) are used to capture the DNA polymerase: when a modified base is present, the biophysical dynamics of its movement changes, creating a unique kinetic signature before, during, and after the base incorporation.
SMRT sequencing can be used to detect modified bases in RNA, including m6A sites. In this case, a reverse transcriptase is used as enzyme with ZMWs to observe the cDNA synthesis in real time. The incorporation of synthetically designed m6A sites leaves a kinetic signature and increases the interpulse duration (IPD).
There are some issues concerning the reading of homonucleotide stretches and the base resolution of m6A therein, due to the stuttering of reverse transcriptase. Secondly, the throughput is too low for transcriptome-wide approaches.
One of the most commonly used platform is the SMRT sequencing technology by Pacific Biosciences.