Brownian Microbarcodes for Bioassays
AssigneeAffymetrix, Inc.
InventorsRandall J. True, Yunqing Ma, Aiguo Zhang, Franklin R. Witney
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Abstract
An encoded microparticle carrying a spatial code is provided; and a set of encoded microparticles are provided with distinguishable spatial codes, wherein the codes comply with a pre-determined coding scheme. Presented are also methods of using the encoded microparticles in various biological assays, such as various multiplex quantitative PCR (real-time PCR) and multiplex chromosomal immunoprecipitation (ChIP) assays.
Claims21
A method of quantifying at least two different nucleic acid targets, which comprises:
- providing a set of encoded microparticles, wherein the set of encoded microparticles comprises one or more different detectable spatial codes, and wherein the encoded microparticles comprise:
- a longest dimension less than 50 microns;
- a plurality of segments, wherein the plurality of segments form a spatial code;
- and
- an outer surface, wherein the outer surface encloses the spatial code, and wherein the spatial code is detectable through the outer surface;
- attaching to the set of encoded microparticles a set of nucleic acid probes, wherein each differently coded microparticle has attached thereto probes of a single nucleotide sequence which is complementary to a single target;
- providing a sample comprising or suspected of comprising at least two different nucleic acid targets, wherein each of the two different nucleic acid targets comprises a different nucleic acid sequence;
- incubating the set of encoded microparticles with the sample, a polymerase, double-stranded DNA dye, and amplification primers sufficient to allow PCR to occur of the two or more target nucleic acids, under conditions which allow polymerization of the two or more target nucleic acid;
- and
- detecting in real time appearance of dye signal for each differently coded encoded microparticle.
A method of determining the sequence of one or more protein-bound subgenomic nucleic acids, which comprises:
- providing a sample comprising cells;
- cross-linking one or more DNA binding proteins to genomic DNA in the sample;
- lysing the cells in the sample;
- shearing the genomic DNA in the sample;
- incubating the sample with antibodies, wherein the antibodies are specific for DNA one or more binding proteins;
- isolating the antibody-bound proteins;
- incubating the antibody-bound proteins with a set of one or more differently coded encoded microparticles, wherein the encoded microparticles have attached thereto unique probe sequences which are complementary to the DNA-binding protein bound nucleic acids, and wherein the encoded microparticles comprise:
- a longest dimension less than 50 microns;
- a plurality of segments, wherein the plurality of segments form a spatial code;
- and
- an outer surface, wherein the outer surface encloses the spatial code, and wherein the spatial code is detectable through the outer surface;
- incubating the set of one or more differently coded encoded microparticles with a label amplification system;
- detecting the label;
- detecting the codes of the one or more differently coded encoded microparticles;
- and
- correlating the encoded microparticle codes with the label signals to determine the sequence of the DNA-binding protein bound one or more subgenomic nucleic acids.
Dependencies are inferred from the claim text.