Single-molecule denaturation mapping of DNA in nanofluidic channels

TY - JOUR

T1 - Single-molecule denaturation mapping of DNA in nanofluidic channels

AU - Reisner, Walter

AU - Larsen, Niels B

AU - Silahtaroglu, Asli

AU - Kristensen, Anders

AU - Tommerup, Niels

AU - Tegenfeldt, Jonas O

AU - Flyvbjerg, Henrik

N1 - Keywords: Algorithms; Bacteriophages; Benzoxazoles; DNA; Formamides; Microfluidic Analytical Techniques; Models, Chemical; Nanotechnology; Nucleic Acid Conformation; Nucleic Acid Denaturation; Quinolinium Compounds; Transition Temperature

PY - 2010/7/27

Y1 - 2010/7/27

N2 - 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.

AB - 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.

U2 - 10.1073/pnas.1007081107

DO - 10.1073/pnas.1007081107

M3 - Journal article

C2 - 20616076

SN - 0027-8424

VL - 107

SP - 13294

EP - 13299

JO - Proceedings of the National Academy of Sciences of the United States of America

JF - Proceedings of the National Academy of Sciences of the United States of America

IS - 30

ER -