Detection of nucleic acid reactions on bead arrays
원문보기
IPC분류정보
국가/구분
United States(US) Patent
등록
국제특허분류(IPC7판)
C12Q-001/68
C12M-001/36
C12P-019/34
C07H-021/04
출원번호
US-0913922
(2013-06-10)
등록번호
US-9279148
(2016-03-08)
발명자
/ 주소
Gunderson, Kevin
Stuelpnagel, John R.
Chee, Mark S.
Fan, Jian-Bing
출원인 / 주소
Illumina, Inc.
대리인 / 주소
Knobbe, Martens, Olson & Bear LLP
인용정보
피인용 횟수 :
0인용 특허 :
254
초록▼
The present invention is directed to methods and compositions for the use of micro sphere arrays to detect and quantify a number of nucleic acid reactions. The invention finds use in genotyping, i.e. the determination of the sequence of nucleic acids, particularly alterations such as nucleotide subs
The present invention is directed to methods and compositions for the use of micro sphere arrays to detect and quantify a number of nucleic acid reactions. The invention finds use in genotyping, i.e. the determination of the sequence of nucleic acids, particularly alterations such as nucleotide substitutions (mismatches) and single nucleotide polymorphisms (SNPs). Similarly, the invention finds use in the detection and quantification of a nucleic acid target using a variety of amplification techniques, including both signal amplification and target amplification. The methods and compositions of the invention can be used in nucleic acid sequencing reactions as well. All applications can include the use of adapter sequences to allow for universal arrays.
대표청구항▼
1. A method of detecting a plurality of target nucleic acid sequences, comprising: a) providing a plurality of target sequences that are immobilized to one or more first solid supports;b) hybridizing a plurality of different first primers to first portions of said plurality of target sequences, wher
1. A method of detecting a plurality of target nucleic acid sequences, comprising: a) providing a plurality of target sequences that are immobilized to one or more first solid supports;b) hybridizing a plurality of different first primers to first portions of said plurality of target sequences, wherein each of said different first primers comprises an adapter sequence exogenous to said target sequences;c) hybridizing a plurality of different second primers to second portions of said plurality of target sequences;d) extending said first or said second primers, and then ligating said first and second primers together to form a plurality of different modified primers comprising said adapter sequences;e) amplifying said plurality of different modified primers to form amplified products comprising said adapter sequences;f) hybridizing said adapter sequences of said amplified products, or their complements, to an array of capture probes attached to a second solid support, thereby forming hybridized capture probes;g) modifying said hybridized capture probes by polymerase extension to form modified capture probes, andh) detecting said modified capture probes, thereby detecting said plurality of target nucleic acid sequences. 2. The method of claim 1, wherein said amplifying comprises hybridizing said plurality of different modified primers with a plurality of amplifier probes complementary to said plurality of first and second primers and amplifying said different modified primers. 3. The method of claim 2, wherein said amplifying comprises a polymerase chain reaction. 4. The method of claim 1, further comprising identifying a nucleotide at a detection position for each of said target nucleic acid sequences, wherein a primer of said plurality of first primers or said plurality of second primers is complementary to said detection position. 5. The method of claim 1, wherein said array comprises a population of beads comprising said capture probes. 6. The method of claim 5, wherein said beads are associated with individual sites of said second solid support. 7. The method of claim 6, wherein each of said sites is configured to have a single associated bead. 8. The method of claim 6, wherein said solid support comprises a fiber optic bundle. 9. The method of claim 1, wherein said array is made by a method selected from the group consisting of a spotting technique, photolithographic technique, and printing technique. 10. The method of claim 1, wherein detecting said modified capture probes comprises detecting a label attached to said modified capture probes. 11. The method of claim 10, wherein detecting said modified capture probes comprises detecting a fluorescent label. 12. The method of claim 1, wherein said plurality of target sequences is immobilized by covalent attachment to said one or more first solid supports. 13. The method of claim 1, wherein said capture probes are covalently attached to said second solid support. 14. The method of claim 1, further comprising a wash step prior to said amplifying of said different modified primers. 15. The method of claim 1, wherein said target sequences comprise loci having a single nucleotide polymorphism (SNP) allele. 16. The method of claim 15, wherein said plurality of said first primers comprise allele specific primers and said plurality of second primers comprise locus specific primers. 17. The method of claims 16, wherein the terminal base of said allele specific primers correspond to said SNP allele. 18. The method of claim 16, further comprising a plurality of second allele specific primers, wherein said plurality of allele specific primers comprises a match for said SNP and said plurality of second allele specific primers comprises a mismatch for said SNP at said detection position. 19. The method of claims 16, wherein step (d) comprises extending said allele specific primer to add a base to basepair with said SNP. 20. The method of claim 16, wherein said detecting indicates a SNP clinically related to a phenotypic variant. 21. The method of claim 16, wherein said detecting determines the genotype of a target sequence that is an indicator of a disease. 22. The method of claim 1, wherein said detecting further comprises determining the amount of a target sequence present in said plurality of nucleic acid sequences. 23. The method of claim 1, wherein said detecting comprises determining the presence of a target sequence that is an indicator of a disease. 24. The method of claim 1, wherein said first and second primers are separated by one or more bases.
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이 특허에 인용된 특허 (254)
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