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Combinatorial decoding of random nucleic acid arrays

  • US 9,163,283 B2
  • Filed: 09/24/2013
  • Issued: 10/20/2015
  • Est. Priority Date: 05/20/1999
  • Status: Active Grant
First Claim
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1. A method of determining a nucleotide sequence of a captured nucleic acid, said method comprising the steps of:

  • a) providing a substrate comprising a first subpopulation and a second subpopulation of captured nucleic acids distributed thereon, wherein captured nucleic acids of the first subpopulation and captured nucleic acids of the second subpopulation comprise an anchor sequence adjacent to a target sequence, the anchor sequence of the first subpopulation of captured nucleic acids having the same sequence as the anchor sequence of the second subpopulation of captured nucleic acids and the target sequence of the first subpopulation of captured nucleic acids being different from the target sequence of the second subpopulation of captured nucleic acids,b) hybridizing a first set of target probes to captured nucleic acids of the first subpopulation and captured nucleic acids of the second subpopulation, said first set of target probes comprising a first variable region, said first variable region comprising at least one detection nucleotide at a detection position, said detection nucleotide being able to basepair with a nucleotide of at least one of said different target sequences, wherein said first set of target probes comprises different groups, and wherein target probes from different groups comprise a different detection nucleotide at the same detection position and a label that distinguishes the detection nucleotide;

    c) detecting the presence of said label;

    d) associating the label with the identity of one or more nucleotides of at least one of said different target sequences;

    e) dehybridizing said target probes from said captured nucleic acids;

    f) hybridizing a second set of target probes to said first and second subpopulations of captured nucleic acids, wherein said second set of probes comprises a second variable region, said second variable region comprising a second detection nucleotide at a detection position, said second detection nucleotide being able to basepair with a nucleotide of at least one of said different target sequences, wherein said plurality of target probes from said second set comprises different groups, and wherein target probes from different groups of said second set comprise a different detection nucleotide at the same detection position and a label that distinguishes the second detection nucleotide, andg) detecting the presence of a label in said second set of target probes.

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