Angewandte Chemie International Edition

Cover image for Vol. 53 Issue 28

Editor: Peter Gölitz, Deputy Editors: Neville Compton, Haymo Ross

Online ISSN: 1521-3773

Associated Title(s): Angewandte Chemie, Chemistry - A European Journal, Chemistry – An Asian Journal, Zeitschrift für Chemie

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Press Release

Angewandte Chemie International Edition ,
doi: 10.1002/anie.201300413

Nr. 15/2013
April 24, 2013

Threaded through a Pore

Single-molecule detection of hydroxymethylcytosine in DNA

Contact: Hagan Bayley, University of Oxford (UK)
Registered journalists may download the original article here:
Single-Molecule Detection of 5-Hydroxymethylcytosine in DNA through Chemical Modification and Nanopore Analysis

Changes in the bases that make up DNA act as markers, telling a cell which genes it should read and which it shouldn’t. In the journal Angewandte Chemie, a British team has now introduced a new method that makes it possible to enrich the rare gene segments that contain the modified base hydroxymethylcytosine and to identify individual hydroxymethylcytosine molecules in DNA. Such modifications are associated with autoimmune diseases and cancer.

Threaded through a Pore - Single-molecule detection of hydroxymethylcytosine in DNA
© Wiley-VCH

The bases adenine, guanine, cytosine, and thymine make up the genetic code. Every cell of the body contains an identical set of complete genetic material. However, the various tissues in the body are very different from each other. This is because the cells have the ability to transcribe only a specific selection of genes into proteins, leaving other genes unused. Epigenetic factors such as “markers” on the DNA control this process. The base cytosine can be equipped with different side groups, such as a methyl or hydroxymethyl group. Dense methylation of regulatory gene segments switches off the corresponding genes. During development of the embryo, methylation patterns initiate cell differentiation. Changes in the methylation patterns are associated with autoimmune diseases and cancer. Hydroxymethylcytosine patterns also seem to play a role in the differentiation of embryonic stem cells as well as in gene expression in cells of the central nervous system.

Sequencing techniques that can be used to specifically detect epigenetic bases are thus very important. To date, the identification of hydroxymethylcytosine has required complex, expensive, or error-prone processes. A team led by Hagan Bayley at the University of Oxford University has now developed a chemical modification that allows for the differentiation of hydroxymethylcytosine and methylcytosine through sequencing in nanopores.

Developed by Oxford Nanopore, a company formed by Hagan Bayley in 2005, the nanopore method is a highly promising alternative to the sequencing of individual DNA molecules without an amplification step. Fed by an enzyme, a single strand of DNA threads through a membrane-embedded protein pore. Depending on which of the bases is in the narrowest part of the pore at a given time, there is a characteristic change in the flow of current through the pore.

A chemical reaction between hydroxymethylcytosine, bisulfite, and a cysteine-containing peptide that leaves the other bases—including methylcytosine—unchanged, greatly improves the resolution as the various bases result in differences in current.

Importantly, it is possible to attach a fluorescent marker to the modified site, or a molecular “eye” that can be used to attach the rare hydroxymethylcytosine-containing DNA fragments to “hooks” that allow the fragments to be enriched over unmodified fragments, enabling rapid sequence analysis.

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About the Author

Hagan Bayley is the Professor of Chemical Biology at the University of Oxford, and the founder of Oxford Nanopore Technologies. He was recognized by the RSC as Chemistry World Entrepreneur of the Year in 2009 and was the society's Interdisciplinary Prize winner in 2012. He is a Fellow of the Royal Society.

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