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US 9,809,844 B2Granted
Method of DNA analysis using micro/nanochannel
AssigneeWisconsin Alumni Research Foundation
InventorsDavid C. Schwartz, Kyubong Jo, Dalia M. Dhingra
Life of this patent
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Abstract
Methods are provided for tagging, characterizing and sorting double-stranded biomolecules while maintaining the integrity of the biomolecules.
Claims8
A method of preparing a double-stranded DNA molecule for optical analysis, the method comprising the steps of:
- ligating the DNA molecule to remove inherent nicks;
- nicking at least one predefined sequence position of one strand of the DNA molecule without breaking the double-stranded DNA molecule into wholly separate pieces, wherein at least one nicked position defines a pattern-characteristic of the nucleotide sequence of the DNA molecule;
- elongating the DNA molecule in a buffer having an ionic strength sufficiently low to increase the persistence length of the DNA molecule;
- labeling the DNA molecule with at least one fluorescent label wherein the fluorescent label is specific to the nicked position;
- and
- introducing the elongated and labeled DNA molecule suspended in the buffer into a channel to maintain the elongated and labeled DNA molecule in an elongated state capable of being analyzed optically without fixation to the channel.
3 more elements
A method of providing an elongated and labeled DNA molecule in a channel for optical analysis, the method comprising:
- (a) providing the DNA molecule in a buffer having an ionic strength sufficiently low that the molecule adopts an elongated form;
- (b) labeling the DNA molecule with at least one fluorescent label;
- and
- (c) introducing the elongated and labeled DNA molecule provided in the buffer into a channel having a cross-sectional dimension that constrains the elongated and labeled DNA which is at least as wide as the elongated and labeled DNA molecule and is less than 1000 nanometers, whereby the elongated DNA is constrained in the channel in the focal plane of a microscope objective, and wherein the elongated DNA molecule remains in the elongated state suitable for being analyzed optically without fixation to the channel.
1 more element
Dependencies are inferred from the claim text.