초록 ▼

A method for detecting nucleic acids by (a) providing a sample having target nucleic acids, each nucleic acid having contiguous first, second, and third domains; (b) contacting the sample with probe sets to form hybridization complexes, wherein each probe set includes (i) a first probe having a sequ

A method for detecting nucleic acids by (a) providing a sample having target nucleic acids, each nucleic acid having contiguous first, second, and third domains; (b) contacting the sample with probe sets to form hybridization complexes, wherein each probe set includes (i) a first probe having a sequence that is complementary to the first domain; and (ii) a second probe having a sequence substantially complementary to the third domain; (c) extending the first probes along the second domains of the complexes while the complexes are immobilized on a solid support; (d) ligating the extended first probes to the second probes to form templates; (e) amplifying the templates with primers that are complementary to the first and second priming sequences to produce amplicons; and (f) detecting the amplicons on the surface of a nucleic acid array.

대표청구항 ▼

1. A method for detecting different target nucleic acids comprising DNA in a sample, each nucleic acid comprising, from 3′ to 5′: contiguous first, second, and third target domains, the method comprising: (a) providing a sample having the different target nucleic acids comprising DNA;(b) contacting

1. A method for detecting different target nucleic acids comprising DNA in a sample, each nucleic acid comprising, from 3′ to 5′: contiguous first, second, and third target domains, the method comprising: (a) providing a sample having the different target nucleic acids comprising DNA;(b) contacting the sample with a plurality of at least 100 different probe sets to form hybridization complexes with the different target nucleic acids, wherein each probe set comprises: (i) a first probe comprising, from 5′ to 3′: a first priming sequence and a sequence that is substantially complementary to the first target domain; and(ii) a second probe comprising 5′ to 3′: a sequence substantially complementary to the third target domain, and a second priming sequence,wherein at least one probe in each of the different probe sets contains a distinct adapter sequence not native to the target nucleic acid;(c) contacting the hybridization complexes with an extension enzyme and nucleotides, wherein the first probes are extended along the second target domains of hybridization complexes formed in (b), wherein the hybridization complexes are immobilized on a solid support when contacted with the extension enzyme and the nucleotides;(d) ligating the extended first probes to the second probes to form amplification templates;(e) amplifying the amplification templates with first and second primers that are complementary to the first priming sequence and the second priming sequence to produce amplicons; and(f) detecting the amplicons on the surface of a nucleic acid array that is different from the solid support that is immobilized to the hybridization complexes. 2. The method of claim 1, wherein the sample is contacted with a plurality of at least 100,000 different probe sets in (b). 3. The method of claim 1, wherein the solid support comprises a plurality of beads. 4. The method of claim 1, wherein the solid support comprises a first binding partner capable of binding to a second binding partner, and the target nucleic acids comprise the second binding partner. 5. The method of claim 4, wherein the first binding partner is streptavidin. 6. The method of claim 1, wherein the first and second primers are universal primers. 7. The method of claim 1, wherein the amplicons are detected on the surface of the nucleic acid array by sequencing. 8. The method of claim 7, wherein the sequencing is pyrosequencing. 9. The method of claim 1, wherein the first probe in each of the sets contains the distinct adapter sequence that is not native to the target nucleic acid. 10. The method of claim 1, wherein the second probe in each of the sets contains the distinct adapter sequence that is not native to the target nucleic acid. 11. The method of claim 1, wherein the different target nucleic acids comprise genomic DNA. 12. The method of claim 1, wherein the sample is a human sample. 13. The method of claim 1, wherein each of the amplicons comprises the distinct adapter sequence of the at least one probe in each of the different probe sets. 14. The method of claim 13, wherein the distinct adapter sequence of each amplicon is hybridized to a capture probe on the surface of the nucleic acid array. 15. The method of claim 1, wherein the detecting of the amplicons on the surface of a nucleic acid array comprises detecting fluorescent labels. 16. The method of claim 1, wherein the distinct adapter sequence of each amplicon is detected on the surface of the nucleic acid array. 17. The method of claim 1, wherein the first probe is extended along the second target domain by addition of a single nucleotide. 18. The method of claim 1, wherein the first probe is extended along the second target domain by addition of multiple nucleotides. 19. A method for detecting different target nucleic acids in a plurality of samples, comprising performing the steps of claim 1 on a plurality of different samples. 20. The method of claim 19, wherein the steps are performed on a plurality of different samples simultaneously.