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MULTIPLEX NUCLEIC ACID REACTIONS

  • US 20130244882A1
  • Filed: 09/06/2012
  • Published: 09/19/2013
  • Est. Priority Date: 02/07/2000
  • Status: Active Grant
First Claim
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1. A method for detecting different target nucleic acids in a sample, each nucleic acid comprising, from 3′

  • to 5′

    ;

    contiguous first, second, and third target domains, comprising;

    (a) providing a sample having different target nucleic acids;

    (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.

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