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Single-molecule denaturation mapping of DNA in nanofluidic channels.

ABSTRACT: Here we explore the potential power of denaturation mapping as a single-molecule technique. By partially denaturing YOYO-1-labeled DNA in nanofluidic channels with a combination of formamide and local heating, we obtain a sequence-dependent "barcode" corresponding to a series of local dips and peaks in the intensity trace along the extended molecule. We demonstrate that this structure arises from the physics of local denaturation: statistical mechanical calculations of sequence-dependent melting probability can predict the barcode to be observed experimentally for a given sequence. Consequently, the technique is sensitive to sequence variation without requiring enzymatic labeling or a restriction step. This technique may serve as the basis for a new mapping technology ideally suited for investigating the long-range structure of entire genomes extracted from single cells.

SUBMITTER: Reisner W

PROVIDER: S-EPMC2922186 | biostudies-literature | 2010 Jul

REPOSITORIES: biostudies-literature

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Single-molecule denaturation mapping of DNA in nanofluidic channels.

Reisner Walter W Larsen Niels B NB Silahtaroglu Asli A Kristensen Anders A Tommerup Niels N Tegenfeldt Jonas O JO Flyvbjerg Henrik H

Proceedings of the National Academy of Sciences of the United States of America 20100707 30

Here we explore the potential power of denaturation mapping as a single-molecule technique. By partially denaturing YOYO-1-labeled DNA in nanofluidic channels with a combination of formamide and local heating, we obtain a sequence-dependent "barcode" corresponding to a series of local dips and peaks in the intensity trace along the extended molecule. We demonstrate that this structure arises from the physics of local denaturation: statistical mechanical calculations of sequence-dependent melting ...[more]

PMID: 20616076

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