Detection of nucleic acid reactions on bead arrays
First Claim
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.
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Abstract
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.
279 Citations
24 Claims
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1. A method of detecting a plurality of target nucleic acid sequences, comprising:
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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, and h) detecting said modified capture probes, thereby detecting said plurality of target nucleic acid sequences. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24)
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Specification