The present disclosure provides methods, compositions and kits as well as systems for manipulating nucleic acids, including implementing isothermal amplification, such as recombinase-polymerase amplification (RPA), of a nucleic acid template using a pre-seeded solid support. Provided are rapid and efficient methods for generating template nucleic acid molecules comprising specific nucleotide sequence bound to solid support. Such methods can be used, for example, in manipulating nucleic acids in preparation for analysis methods that utilize monoclonal populations of nucleic acids.

Various embodiments of a low-volume sequencing system are provided herein. The system can include a low-volume flowcell having at least one reaction chamber of a defined volume (e.g., less than about 100 l). The system can also include an automated reagent delivery mechanism configured to reversibly couple with the inlet port corresponding to a target reaction chamber thereby placing allowing for reagent to be accurately moved from a storage container to the reaction chamber with minimal reagent waste. The flowcells can include a plurality of reaction chambers (e.g., 6) thereby allowing for parallel analysis of multiple samples. Various methods of analyzing a biomolecule are also provided herein.

According to various embodiments, a method is provided that comprises washing an array of DNA-coated beads on a substrate, with a wash solution to remove stacked beads from the substrate. The wash solution can include inert solid beads in a carrier. The DNA-coated beads can have an average diameter and the solid beads in the wash solution can have an average diameter that is at least twice the diameter of the DNA-coated beads. The washing can form dislodged DNA-coated beads and a monolayer of DNA-coated beads. In some embodiments, first beads for forming an array are contacted with a poly(ethylene glycol) (PEG) solution comprising a PEG having a molecular weight of about 350 Da or less. In some embodiments, slides for forming bead arrays are provided as are systems for imaging the same.

This invention provides a technique enabling to detect target molecules of low concentration with high sensitivity. This invention includes (i) a step of introducing a hydrophilic solvent (42) containing beads (40),(41) into a space (30) between (a) a lower layer section (10) including a plurality of receptacles (13) each of which is capable of storing only one of the beads (41),(41) and which are separated from each other by a side wall (12) having a hydrophobic upper surface and (b) an upper layer section (20) facing a surface of the lower layer section (10) on which surface the plurality of receptacles (13) are provided; and (ii) a step of introducing a hydrophobic solvent (43) into the space (30), the step (ii) being carried out after the step (i).

This invention provides a technique enabling to detect target molecules of low concentration with high sensitivity. This invention includes (i) a step of introducing a hydrophilic solvent (42) containing beads (40),(41) into a space (30) between (a) a lower layer section (10) including a plurality of receptacles (13) each of which is capable of storing only one of the beads (41),(41) and which are separated from each other by a side wall (12) having a hydrophobic upper surface and (b) an upper layer section (20) facing a surface of the lower layer section (10) on which surface the plurality of receptacles (13) are provided; and (ii) a step of introducing a hydrophobic solvent (43) into the space (30), the step (ii) being carried out after the step (i).

A microsphere-based analytic chemistry system and method for making the same is disclosed in which microspheres or particles carrying bioactive agents may be combined randomly or in ordered fashion and dispersed on a substrate to form an array while maintaining the ability to identify the location of bioactive agents and particles within the array using an optically interrogatable, optical signature encoding scheme. A wide variety of modified substrates may be employed which provide either discrete or non-discrete sites for accommodating the microspheres in either random or patterned distributions. The substrates may be constructed from a variety of materials to form either two-dimensional or three-dimensional configurations. In a preferred embodiment, a modified fiber optic bundle or array is employed as a substrate to produce a high density array. The disclosed system and method have utility for detecting target analytes and screening large libraries of bioactive agents.

The invention provides particle compositions having applications in nucleic acid analysis. Nucleic acid polymer particles of the invention allow polynucleotides to be attached throughout their volumes for higher loading capacities than those achievable solely with surface attachment. In one aspect, nucleic acid polymer particles of the invention comprise polyacrylamide particles with uniform size distributions having low coefficients of variations, which result in reduced particle-to-particle variation in analytical assays. Such particle compositions are used in various amplification reactions to make amplicon libraries from nucleic acid fragment libraries.

Complete nucleic acid library preparation devices are provided. Aspects of the devices include: a thermal chip module comprising multiple CLC reaction wells; one or more plate locations; a robotically controlled liquid handler configured to transfer liquid between the one or more plate locations and the thermal chip module; and a bulk reagent dispenser configured to access each CLC reaction well of the thermal chip module.

A method for determining a presence or absence of one or more target analytes in a sample includes contacting the sample with an array of particles comprising at least first and second particle subsets disposed therein with a known particle number ratio with respect to each other. The first particle subset has at least one binding site configured to bind with a first target analyte and the second particle subset has at least one binding site configured to bind with a second, different target analyte. Changes are detected in a detectable signal emitted by the particles after contacting the sample with the array. A number of the particles that emit the change in the detectable signal are counted and this number is compared to the known particle number ratio of the subsets so as to determine the presence or absence of the one or more of the target analytes.

This invention provides high unit density arrays of microparticles and methods of assembling such arrays. The microparticles in the arrays may be functionalized with chemical or biological entities specific to a given target analyte. The high unit density arrays of this invention are formed on chips which may be combined to form multichip arrays according to the methods described herein. The chips and/or multichip arrays of this invention are useful for chemical and biological assays.