A single-molecule barcoding system using nanoslits for DNA analysis.
Molecular confinement offers new routes for arraying large DNA molecules, enabling single-molecule schemes aimed at the acquisition of sequence information. Such schemes can rapidly advance to become platforms capable of genome analysis if elements of a nascent system can be integrated at an early stage of development. Integrated strategies are needed for surmounting the stringent experimental requirements of nanoscale devices regarding fabrication, sample loading, biochemical labeling, and detection. We demonstrate that disposable devices featuring both micro- and nanoscale features can greatly elongate DNA molecules when buffer conditions are controlled to alter DNA stiffness. Furthermore, we present analytical calculations that describe this elongation. We also developed a complementary enzymatic labeling scheme that tags specific sequences on elongated molecules within described nanoslit devices that are imaged via fluorescence resonance energy transfer. Collectively, these developments enable scaleable molecular confinement approaches for genome analysis.
Jo
Kyubong
K
Laboratory for Molecular and Computational Genomics, University of Wisconsin, 425 Henry Mall, Madison, WI 53706, USA.
Dhingra
Dalia M
DM
Odijk
Theo
T
de Pablo
Juan J
JJ
Graham
Michael D
MD
Runnheim
Rod
R
Forrest
Dan
D
Schwartz
David C
DC
D001704:Biopolymers / Q000032:analysis / Q000737:chemistry; D002021:Buffers; D022202:Chromosomes, Artificial, Bacterial / Q000737:chemistry; D004247:DNA / Q000032:analysis* / Q000737:chemistry; D004269:DNA, Bacterial / Q000032:analysis / Q000737:chemistry; D004279:DNA, Viral / Q000032:analysis / Q000737:chemistry; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes; D036103:Nanotechnology; D009690:Nucleic Acid Conformation; D009994:Osmolar Concentration
D001704:Biopolymers; D002021:Buffers; D004269:DNA, Bacterial; D004279:DNA, Viral; D005456:Fluorescent Dyes; D004247:DNA