A University of Cambridge and Cambridge Epigenetix team outlines a sequencing strategy for profiling 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) epigenetic marks in concert with whole-genome sequencing. As they report in Nature Biotechnology, the researchers relied on an enzymatic DNA processing approach for their six-letter sequencing method, which is designed to pick up 5mC and 5hmC as well as all four canonical DNA bases on existing DNA sequencing platforms. After testing a five-letter version of the approach, used to sequence four DNA bases and modified cytosines (5mC and 5hmC) across the genome in a B lymphoblast human cell line on the Illumina Novaseq platform, they tweaked the approach to distinguish between 5mC and 5hmC using DNA methyltransferase 5 and beta-glycosyltransferase enzymes to assess 5mC and protect 5hmC through glycosylation, respectively. The authors found that the six-letter sequencing strategy unearthed 5mC marks with more than 95 percent accuracy in aligned reads, while finding nearly 98 percent of 5hmC bases and offering some 99.9 percent accuracy when it came to sequences for unmodified cytosine bases. "As the platform fundamentally uses Watson-Crick base pairing to decode information," they write, "it can be made readily compatible with any sequencer platform and we see opportunities for its future application to long-read sequencing and single-cell analysis."
UK Team Presents Genetic, Epigenetic Sequencing Method
Feb 06, 2023