DNA sequencing with reagent recycling on wiregrid
AssigneeKoninklijke Philips N.V.
InventorsReinhold Wimberger-Friedl, Johan Lub, Pieter Jan Van Der Zaag
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Abstract
The present invention relates to DNA sequencing with reagent cycling on the wiregrid. The sequencing approach suggested with which allows to use a single fluid with no washing steps. Based on strong optical confinement and of excitation light and of cleavage light, the sequencing reaction can be read-out without washing the surface. Stepwise sequencing is achieved by using nucleotides with optically cleavable blocking moieties. After read-out the built in nucleotide is deblocked by cleavage light through the same substrate. This ensures that only bound nucleotides will be unblocked.
Claims16
A device for optically controlling an iterative stepwise reaction to determine a sequence of a nucleic acid by synthesis, the device comprising:
- a substrate for binding at least one molecule on a first surface of the substrate;
- and
- an optical arrangement comprising a filter disposed on said first surface,
- wherein the optical arrangement is configured to direct excitation light of at least a first excitation wavelength λEx1 to the substrate to excite a fluorescent label of a first nucleotide,
- wherein the first nucleotide is incorporated into the molecule bound on the first surface of the substrate,
- wherein the optical arrangement is configured to receive and detect fluorescent light emitted by the fluorescent label of the first nucleotide,
- wherein the optical arrangement is configured to direct cleavage light of a cleavage wavelength λCL to the substrate to optically induce a cleavage reaction at the first nucleotide to cleave a blocking moiety and the fluorescent label away from the first nucleotide,
- wherein the substrate is configured to provide for an evanescent wave of the excitation light at the first surface of the substrate,
- wherein at least one of the excitation light or the cleavage light is polarized light,
- wherein said filter is configured to reflect said at least one of the excitation light or the cleavage light and to transmit light having a polarization that is different from a polarization of said at least one of the excitation light or the cleavage light.
A method for optically controlling an iterative stepwise reaction to determine a sequence of a nucleic acid by synthesis, the method comprising:
- providing a substrate with a molecule bound on a first surface of the substrate, wherein a filter is disposed on said first surface;
- irradiating the substrate with excitation light of at least a first excitation wavelength λEx1 by an optical arrangement and thereby optically exciting a fluorescent label of a first nucleotide, wherein the first nucleotide is incorporated in the bound molecule on the substrate;
- providing an evanescent wave of the excitation light by the substrate at the first surface of the substrate;
- receiving and detecting fluorescence of the excited fluorescent label of the first nucleotide by the optical arrangement;
- irradiating the substrate with cleavage light of a cleavage wavelength λCL by the optical arrangement and thereby optically inducing a cleaving reaction at the first incorporated nucleotide, wherein at least one of the excitation light or the cleavage light is polarized light;
- and
- reflecting said at least one of the excitation light or the cleavage light by said filter, wherein the filter is configured to transmit light having a polarization that is different from a polarization of said at least one of the excitation light or the cleavage light.
Dependencies are inferred from the claim text.