Abstract: Cyclone Rolling Circle Amplification (CRCA) based next-generation sequencing (NGS) using a multiple primer rolling circle amplification (RCA) reaction is generated using two or more tandem primers from the same strand on different locations of library adaptor regions of double stranded enriched targeted polymerase chain reaction (PCR) library products. This process allows multiple initiation and syntheses of the RCA reaction by an enzyme on the same circular template molecule, which is beneficial since the two or more primers complement each other in generating uniform amplification of the target circle population. Also, a method for keeping DNA nanoballs (also known as rolonies) compact, or more compact, and for pre-priming rolonies before sequencing to eliminate the hybridization of a seqeuncing primer after seeding the rolonies, and a Rolonies rolling circle amplification (RCA) based next-generation sequencing (NGS) using a dual Rolonies primer approach named REPLI-Rolony.

Abstract: The invention relates to methods, compositions, devices, systems and kits as described including, without limitation, reagents and mixtures for determining the identity of nucleic acids in nucleotide sequences using, for example, sequencing by synthesis methods. In particular, the present invention contemplates the use of photoprotective mixture of compounds as imaging reagents to improve stability and storage of fluorescent compounds, including but not limited to, nucleotides with fluorescent labels.

Abstract: The invention relates to methods, compositions, devices, systems and kits as described including, without limitation, reagents and mixtures for determining the identity of nucleic acids in nucleotide sequences using, for example, sequencing by synthesis methods. In particular, the present invention contemplates the use of polyphenolic compounds, known as antioxidant additives, to improve the efficiency of Sequencing-By-Synthesis reactions. For example, gallic acid (GA) is shown herein to be one of many exemplary SBS polyphenolic additives.

Abstract: The invention relates to methods, compositions, devices, systems and kits as described including, without limitation, reagents and mixtures for determining the identity of nucleic acids in nucleotide sequences using, for example, sequencing by synthesis methods. In particular, the present invention contemplates the use of photoprotective mixture of compounds as imaging reagents to improve stability and storage of fluorescent compounds, including but not limited to, nucleotides with fluorescent labels.

Abstract: The invention relates to methods, compositions, devices, systems and kits as described including, without limitation, reagents and mixtures for determining the identity of nucleic acids in nucleotide sequences using, for example, sequencing by synthesis methods. In particular, the present invention contemplates the use of polyphenolic compounds, known as antioxidant additives, to improve the efficiency of Sequencing-By-Synthesis reactions. For example, gallic acid (GA) is shown herein to be one of many exemplary SBS polyphenolic additives.

Abstract: Fiducial marking systems provided for forming a global coordinate system for a visual system having a limited field of view. The fiducial marking system includes a target surface that is movable relative to the visual system and a visual pattern associated with the target surface. The visual pattern includes a plurality of first lines and a plurality of second lines. Each first line is spaced by a first distance from each adjacent first line and each second line is spaced by a second distance from each adjacent second line, wherein the first distance is different than the second distance. Additionally, the first distance and the second distance are selected such that at least one first line and at least one second line are within the field of view at all relative positions of the visual system and the target surface.

Abstract: The invention relates to methods, compositions, devices, systems and kits as described including, without limitation, reagents and mixtures for determining the identity of nucleic acids in nucleotide sequences using, for example, sequencing by synthesis methods. In particular, the present invention contemplates the use of polyphenolic compounds, known as antioxidant additives, to improve the efficiency of Sequencing-By-Synthesis reactions. For example, gallic acid (GA) is shown herein to be one of many exemplary SBS polyphenolic additives.

Abstract: The invention relates to methods, compositions, devices, systems and kits as described including, without limitation, reagents and mixtures for determining the identity of nucleic acids in nucleotide sequences using, for example, sequencing by synthesis methods. In particular, the present invention contemplates the use of polyphenolic compounds, known as antioxidant additives, to improve the efficiency of Sequencing-By-Synthesis reactions. For example, gallic acid (GA) is shown herein to be one of many exemplary SBS polyphenolic additives.

Abstract: A sequencing instrument including a sequencing stage and a microscope. The sequencing stage has a reference plate with a flat reference surface. The microscope optical axis is perpendicular to the reference surface. The sequencing stage is configured to receive a flexible film having a plurality of DNA templates immobilized on a first side of the film, and to hold the flexible film against the flat reference surface with at least some of the plurality of DNA templates in an object plane that is perpendicular to the microscope's optical axis.

Abstract: A flowcell for a sequencing instrument. The flowcell includes a fluid inlet, a fluid outlet, a flow channel formed between an at least partially transparent cover and a base and fluidly connecting the fluid inlet to the fluid outlet, and a capture substrate provided in the flow channel. The capture substrate includes microretainers configured to each receive a single microspot having a microspot diameter, and microretainer is separated from adjacent microretainers by an interstitial gap distance that is equal to or greater than the microspot diameter. A particle separator may be fluidly connected to the flowcell. The particle separator may include a microfluidic channel having an array of micropillars to transfer a plurality of the microspots to a loading buffer that may be delivered to the flowcell.

Abstract: A processing instrument flowcell having a flowcell channel with an upstream channel end, a downstream channel end, a longitudinal axis extending from the upstream channel end to the downstream channel end, and a first operative surface extending between the upstream channel end and the downstream channel end and configured to receive a first number of DNA templates. A first reagent inlet is fluidically connected to the upstream channel end at a location adjacent the first operative surface. A buffer inlet is fluidically connected to the upstream channel end at a location spaced from the first operative surface. An outlet fluidically connected to the downstream channel end. Also provided is a method for operating a flowcell channel under laminar flow conditions to maintain a first reagent adjacent a first operative surface and a buffer fluid spaced from the first operative surface. The flowcell channel may have multiple separate operative surfaces.

Abstract: The invention relates to methods, compositions, devices, systems and kits as described including, without limitation, reagents and mixtures for determining the identity of nucleic acids in nucleotide sequences using, for example, sequencing by synthesis methods. In particular, the present invention contemplates the use of polyphenolic compounds, known as antioxidant additives, to improve the efficiency of Sequencing-By-Synthesis reactions. For example, gallic acid (GA) is shown herein to be one of many exemplary SBS polyphenolic additives.

Abstract: A sequencing instrument including a sequencing stage and a microscope. The sequencing stage has a reference plate with a flat reference surface. The microscope optical axis is perpendicular to the reference surface. The sequencing stage is configured to receive a flexible film having a plurality of DNA templates immobilized on a first side of the film, and to hold the flexible film against the flat reference surface with at least some of the plurality of DNA templates in an object plane that is perpendicular to the microscope's optical axis.

Abstract: A flowcell for a sequencing instrument. The flowcell includes a fluid inlet, a fluid outlet, a flow channel formed between an at least partially transparent cover and a base and fluidly connecting the fluid inlet to the fluid outlet, and a capture substrate provided in the flow channel. The capture substrate includes microretainers configured to each receive a single microspot having a microspot diameter, and microretainer is separated from adjacent microretainers by an interstitial gap distance that is equal to or greater than the microspot diameter. A particle separator may be fluidly connected to the flowcell. The particle separator may include a microfluidic channel having an array of micropillars to transfer a plurality of the microspots to a loading buffer that may be delivered to the flowcell.