Abstract: Labeled nucleotide analogs comprising at least one avidin protein, at least one dye-labeled compound, and at least one nucleotide compound are provided. The analogs are useful in various fluorescence-based analytical methods, including the analysis of highly multiplexed optical reactions in large numbers at high densities, such as single molecule real time nucleic acid sequencing reactions. The analogs are detectable with high sensitivity at desirable wavelengths. They contain structural components that modulate the interactions of the analogs with DNA polymerase, thus decreasing photodamage and improving the kinetic and other properties of the analogs in sequencing reactions. Also provided are nucleotide and dye-labeled compounds of the subject analogs, as well as intermediates useful in the preparation of the compounds and analogs. Compositions comprising the compounds, methods of synthesis of the intermediates, compounds, and analogs, and mutant DNA polymerases are also provided.

Abstract: A method for identifying a nucleotide in a template nucleic acid by (a) providing a plurality of primer-template nucleic acid hybrids, wherein the primers have an extendable 3? end; (b) contacting the plurality with: (i) blocked nucleotides to produce a first subset of the primer-template nucleic acid hybrids that include a blocked nucleotide at the 3? end, and (ii) a ternary complex inhibitor to produce a second subset of the primer-template nucleic acid hybrids that include a ternary complex inhibitor; (c) forming ternary complexes that each include a polymerase, a primer-template nucleic acid hybrid of the first subset, and a cognate nucleotide; and (d) detecting the ternary complexes, thereby identifying a nucleotide in the template nucleic acid.

Abstract: The present invention provides methods, compositions, and systems for distributing molecules and complexes into reaction sites. In particular, the methods, compositions, and systems of the present invention result in loading of polymerase enzyme complexes into a predetermined number of reaction sites, including nanoscale wells.

Abstract: The present disclosure provides compositions, methods and systems for quantifying target sequences and identifying target sequence variants.

Abstract: Compositions that include polymerases with features for improving entry of nucleotide analogues into active site regions and for coordinating with the nucleotide analogues in the active site region are provided. Methods of making the polymerases and of using the polymerases in sequencing and DNA replication and amplification as well as kinetic models of polymerase activity and computer-implemented methods of using the models are also provided.

Abstract: We describe methods of producing gratings such as Bragg gratings with optical waveguides. Described are the writing and erasing of gratings within SiON waveguides by forming standing waves. Methods, systems, instruments, and devices are described that provide improved transmission of light through such waveguides.

Abstract: Provided are pixels, devices, systems, and methods relating to active pixel sensor in which each active pixel puts out a signal that represents a temporal differential. The output of the active pixel represents a comparison or difference between successive frames. The differential can represent the state of the sensor element in the pixel at the end of one frame compared to the state of the sensor element of the frame, at the end of the prior frame.

Abstract: Methods, devices, and systems for performing intermittent detection during analytical reactions are provided. Such methods facilitate collection of reaction data from disparate reaction times. Further, such methods are useful for reducing photo-induced damage of one or more reactants in an illuminated analytical reaction at a given reaction time. In preferred embodiments, the reaction mixture is subjected to at least one illuminated and non-illuminated period and allowed to proceed such that the time in which the reaction mixture is illuminated is less than a photo-induced damage threshold period.

Abstract: Labeled nucleotide analogs comprising at least one avidin protein, at least one dye-labeled compound, and at least one nucleotide compound are provided. The analogs are useful in various fluorescence-based analytical methods, including the analysis of highly multiplexed optical reactions in large numbers at high densities, such as single molecule real time nucleic acid sequencing reactions. The analogs are detectable with high sensitivity at desirable wavelengths. They contain structural components that modulate the interactions of the analogs with DNA polymerase, thus decreasing photodamage and improving the kinetic and other properties of the analogs in sequencing reactions. Also provided are nucleotide and dye-labeled compounds of the subject analogs, as well as intermediates useful in the preparation of the compounds and analogs. Compositions comprising the compounds, methods of synthesis of the intermediates, compounds, and analogs, and mutant DNA polymerases are also provided.

Abstract: Disclosed herein is a novel method to fabricate magnetic silica nanomembranes using thin polymer cores based on silica deposition and self-wrinkling induced by thermal shrinkage. These micro- and nano-scale structures have vastly enlarged the specific area of silica, thus the magnetic silica nanomembranes can be used for solid phase extraction of nucleic acids. The magnetic silica nanomembranes are suitable for nucleic acid purification and isolation and demonstrated better performance than commercial particles in terms of nucleic acid recovery yield and integrity. In addition, the magnetic silica nanomembranes may have high nucleic acid capacity due to significantly enlarged specific surface area of silica. Methods of use and devices comprising the magnetic silica nanomembranes are also provided herein.

Abstract: Provided are compositions comprising recombinant DNA polymerases that include amino acid substitutions, insertions, deletions, and/or exogenous features that confer modified properties upon the polymerase for enhanced single molecule sequencing or nucleic acid amplification. Such properties include enhanced performance with large nucleotide analogs, increased stability, increased readlength, and improved detection of modified bases, and can also include resistance to photodamage, enhanced metal ion coordination, reduced exonuclease activity, reduced reaction rates at one or more steps of the polymerase kinetic cycle, decreased branching fraction, altered cofactor selectivity, increased yield, increased accuracy, altered speed, increased cosolvent resistance, and the like. Also provided are nucleic acids which encode the polymerases with the aforementioned phenotypes, as well as methods of using such polymerases to make a DNA or to sequence a DNA template.

Abstract: Methods of producing substrates having selected active chemical regions by employing elements of the substrates in assisting the localization of active chemical groups in desired regions of the substrate. The methods may include optical, chemical and/or mechanical processes for the deposition, removal, activation and/or deactivation of chemical groups in selected regions of the substrate to provide selective active regions of the substrate.

Abstract: FRET-labeled compounds are provided for use in analytical reactions. In certain embodiments, FRET-labeled nucleotide analogs are used in place of naturally occurring nucleoside triphosphates or other analogs in analytical reactions comprising nucleic acids, for example, template-directed nucleic acid synthesis, DNA sequencing, RNA sequencing, single-base identification, hybridization, binding assays, and other analytical reactions.

Abstract: Sets of compounds bearing detectably different groups of labels are provided. Typically, different compounds bear different numbers of a single type of label and are thus distinguishable by the amplitude of signal produced by the label. The compounds are assembled from label components and protein cores to facilitate modular production of the compounds. In compounds containing two or more proteins, the proteins are typically covalently linked. Useful sets of compounds include sets of labeled nucleotide analogs, particularly dye-label nucleotide analogs that include tetravalent biotin-binding protein cores.

Abstract: Arrays of integrated analytical devices and their methods for production are provided. The arrays are useful in the analysis of highly multiplexed optical reactions in large numbers at high densities, including biochemical reactions, such as nucleic acid sequencing reactions. The devices allow the highly sensitive discrimination of optical signals using features such as spectra, amplitude, and time resolution, or combinations thereof. The devices include an integrated diffractive beam shaping element that provides for the spatial separation of light emitted from the optical reactions.

Abstract: The present disclosure provides improved methods for generating asymmetrically-tagged nucleic acid constructs, compositions comprising such constructs, and kits and systems for generating such constructs.

Abstract: Sequencing methods, devices, and systems are described. Arrays of nanoscale electronic elements comprising two electrodes separated by an insulating layer are used to provide sequence information about a template nucleic acid in a polymerase-template complex bound proximate to the insulating region. A sequencing reaction mixture comprising nucleotide analogs having impedance labels is introduced to the array of nanoscale electronic elements under conditions of polymerase mediated nucleic acid synthesis. The time sequence of incorporation of nucleotide analogs is determined by identifying the types of labels of the nucleotide analogs that are incorporated into the growing strand using measured impedance.

Abstract: Real time electronic sequencing devices, chips, and systems are described. Arrays of nanoFET devices are used to provide sequence information about a template nucleic acid in a polymerase-template complex bound to the nanoFET. The nanoFET devices typically have a source, a drain and a gate comprising a nanowire. A single polymerase enzyme complex comprising a polymerase enzyme complexed with the template nucleic acid is bound to the gate. The polymerase is bound to the gate non-covalently through a polymeric binding agent that has two strands, each strand interacting with the nanowire such that the polymerase is in a central location between the strands with the polymeric binding agent extending away from the polymerase complex along the nanowire in both directions.

Abstract: Provided herein are systems, devices, and methods for improved optical waveguide transmission and alignment in an analytical system. Waveguides in optical analytical systems can exhibit variable and increasing back reflection of single-wavelength illumination over time, thus limiting their effectiveness and reliability. The systems are also subject to optical interference under conditions that have been used to overcome the back reflection. Novel systems and approaches using broadband illumination light with multiple longitudinal modes have been developed to improve optical transmission and analysis in these systems. Novel systems and approaches for the alignment of a target waveguide device and an optical source are also disclosed.

Abstract: Optical analytical devices and their methods of use are provided. The devices are useful in the analysis of highly multiplexed optical reactions in large numbers at high densities, including biochemical reactions, such as nucleic acid sequencing reactions. The devices include optical waveguides for illumination of the optical reactions. The devices further provide for the efficient coupling of optical excitation energy from the waveguides to the optical reactions. Optical signals emitted from the reactions can thus be measured with high sensitivity and discrimination using features such as spectra, amplitude, and time resolution, or combinations thereof. The devices of the invention are well suited for miniaturization and high throughput.