Nanofabricated Closed Fluidic Channels

DNA Fragment Sizing by Single Molecule Detection in Submicrometer-Sized Closed Fluidic Channels

Jonas Korlach, Mathieu Foquet, Warren Zipfel, Watt W Webb, Harold G Craighead

Cornell University, School of Applied Physics, Clark Hall, Ithaca, New York 14853

The fabrication of fluidic channels with dimensions smaller than 1 mm is described and characterized in respect to their use for detection of individual DNA molecules. The small dimensions facilitate single molecule detection and minimize events of simultaneous passage of more than one molecule through the measurement volume. The behavior of DNA inside these channels under an applied electrical field was first studied using M13 double-stranded DNA. A linear relationship between flow speed and applied electric field across the channel was observed. Flow speeds were measured using Fluorescence Correlation Spectroscopy. Speeds as high as 5 mm/s were reached, corresponding to only a few milliseconds of analysis time per molecule.

The channels were then used to characterize a mixture of 7 DNA fragments. Arrays of Gaussian peaks were used to fit the photon burst histograms. The distribution and relative proportions of the individual fragments, as well as the overall concentration of the DNA sample were deduced from a single experiment. The burst size was linearly proportional to the DNA fragment size over the entire range of the molecular weight marker. The amount of sample required for the analysis was about 10,000 molecules.

References:

Foquet et al. (2002) Anal. Chem. ASAP Article 10.1021/ac011076w.

Supported by NCRR-NIH grant P41-RR04224 and DOE grant DE-FG02-99ER62809.

Movies of YOYO-1 stained DNA flowing in the nanochannel

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