Bioarray Solutions Ltd.

Other

Jiacheng Yang

Michael Seul

Sukanta Banerjee

Chiu Wo Chau

Ye Hong

Tatiana Vener

Hui Huang

Yi Zhang

Enqing Tan

Nataliya Korzheva

Disclosed are methods of multiplexed analysis of oligonucleotides in a sample, including a method of preventing a significant reduction in duplexes detectable in a hybridization assay involving (i) selecting probe lengths for sets of oligonucleotide probes, wherein probes include different subsequences such that at least one subsequence is complementary to a subsequence in a cognate target; wherein probes for longer cognate targets are longer in length than probes for shorter cognate targets, (ii) selecting, for each set of probes, a density of oligonucleotides probes attached per unit area on a solid phase carrier which is below a limit at which the significant reduction in detectable duplexes is predicated to take place, (iii) producing the probes and affixing them to different solid phase carriers at the selected density, and (iv) annealing targets to the probes, wherein signal intensities of probes and targets of different lengths are about the same.

Optimization of gene expression analysis using immobilized capture probes

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.

Arrays of microparticles and methods of preparation thereof

Disclosed herein are methods for array assembly and detection. The methods can use an incubation chamber containing a suspension of nucleic acid targets, polymerase and a set of oligonucleotide probes bound to magnetic beads in a randomly dispersed state. Each probe can have a target binding domain that is complementary to a target nucleic acid, a closing domain with a sequence that is complementary to the sequence of the target binding domain, and a joining region between the binding domain and the closing domain, which is not complementary to the target nucleic acid. Method steps can include providing the incubation chamber, placing the incubation chamber in a magnetic trap, generating a magnetic field that induces the magnetic beads to migrate towards a substrate and, once in proximity to the substrate, to interact with each other repulsively and reorganize into arrays, and imaging the array.

Methods for array assembly and detection involving elongation of self-complementary looped probes

Disclosed are methods of multiplexed analysis of oligonucleotides in a sample, including: methods of probe and target 'engineering', as well as methods of assay signal analysis relating to the modulation of the probe-target affinity constant, K by a variety of factors including the elastic properties of target strands and layers of immobilized ('grafted') probes; and assay methodologies relating to: the tuning of assay signal intensities including dynamic range compression and on-chip signal amplification; the combination of hybridization-mediated and elongation-mediated detection for the quantitative determination of abundance of messages displaying a high degree of sequence similarity, including, for example, the simultaneous determination of the relative expression levels, and identification of the specific class of, untranslated AU-rich subsequences located near the 3' terminus of mRNA; and a new method of subtractive differential gene expression analysis which requires only a single color label.

In a multiplexed assay method carried out in solution, wherein the solution contains nucleic acid targets and, wherein several different types of oligonucleotide probes, each type having a different sequence in a region designated as a target binding domain, are used to detect the nucleic acid targets, said assay method including a method for increasing the effective concentration of the nucleic acid targets at the surface of a bead to which the oligonucleotide probes are bound, by one or more of the following steps: adjusting assay conditions so as to increase the effective concentration of the targets available for binding to the probes, by one or more of the following: (i) selecting a particular probe density on the surface of the bead; (ii) selecting a solution having an ionic strength greater than a threshold; (ii) selecting a target domain of a size less than a threshold; or (iii) selecting target domains within a specified proximity to a terminal end of the targets.

Probe density self-considerations and elongation of complementary looped probes where probes are attached to a solid phase

A method of fragmentation of double stranded DNA is disclosed for use in nucleic acid analysis, notably in the multiplexed analysis of polymorphisms and mutations. The method produces a multiplicity of labeled sense and anti-sense fragments which are not complementary, and thus do not significantly re-anneal under conditions suitable for hybridization analysis (or capture-mediated elongation analysis) of the polymorphisms and/or mutations. The fragments display a desired or predicted length distribution. Cleavage sites can be selected such that the fragments are short, yet long enough to allow discrimination among fragments in an assay, and as a matter of statistical probability, such that the majority of fragments contain at least one labeled nucleotide to facilitate detection.

Kits for multiplexed nucleic acid analysis by capture of single-stranded DNA produced from double-stranded target fragments

Disclosed are methods of for constructing a bead-displayed library of oligonucleotide probes (or sequence-modified capture moieties such as protein-nucleic acid conjugates) by ligation of a capture probe, having an analyte-specific sequence, to an anchor probe that is attached, at its 5'-end, (or possibly at the 3&#x2032; end) to an encoded carrier such as a color-coded microparticle ('bead'). Such a library can also be constructed by elongation of an anchor probe, using a second probe as the elongation template, wherein the second probe has an anchor-specific subsequence and an analyte-specific subsequence.

Creation of functionalized microparticle libraries by oligonucleotide ligation or elongation

A method mediated with in-vitro transcription ('IVT') which permits miniaturization of multiplexed DNA and RNA analysis, and in which elongation-mediated multiplexed analysis of polymorphisms (eMAP&#xae;) is used as the analysis step, is described. Also described is a method mediated with IVT is for selecting a designated strand from T7-tagged double stranded DNA: wherein, the selected strand forms the template for RNA synthesis. In one embodiment, double stranded DNA incorporating the T7 (or other) promoter sequence at the 3' end or the 5&#x2032; end is produced, for example, by amplification of genomic DNA using the Polymerase Chain Reaction (PCR). Also disclosed are nested PCR designs permitting allele analysis in combination with strand selection by IVT. Further, in one embodiment of a homogeneous format for transcription-mediated amplification and multiplexed detection (which may be particularly suited for viral or pathogen detection), encoded microparticles display 'looped' capture probe configurations permitting the generation of a signal upon capture of RNA product and real-time assay monitoring.

Nucleic acid amplification with integrated multiplex detection

The present invention relates to molecular constructs and methods of their use in detecting biochemical reactions. In particular, the invention relates to a molecular construct having a capture portion and a substrate portion, where the capture portion isolates the construct from a sample medium, and the substrate portion enables the construct to be acted upon and undergo a physical change which can be detected and measured. These molecular constructs may be used in diagnostic assays, genetic screening for potential risks of certain diseases in individuals, and drug discovery and toxicogenomics, using high throughput screening of compounds.

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Hillsboro, NJ, United States of America

Jiacheng Yang