Digital counting of individual molecules by stochastic attachment of diverse label-tags

  • US 9,315,857 B2
  • Filed: 12/15/2011
  • Issued: 04/19/2016
  • Est. Priority Date: 12/15/2009
  • Status: Active Grant
First Claim
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1. A method comprising:

  • a) attaching a plurality of diverse label-tags to a nucleic acid target from a sample that contains multiple copies of the nucleic acid target, thereby producing a plurality of labeled targets, wherein;

    i) a label-tag of the plurality of diverse label-tags comprises nucleotides selected from purine bases, pyrimidine bases, natural nucleotide bases, chemically modified nucleotide bases, biochemically modified nucleotide bases, non-natural nucleotide bases; and

    ii) a labeled target of the plurality of labeled targets comprises a distinct label-tag and at least a portion of a nucleic acid target, or its complementary sequence;

    b) amplifying the plurality of labeled-targets to produce a plurality of amplified labeled-targets, wherein an amplified labeled-target of the plurality of amplified labeled-targets comprises a copy of at least a portion of the nucleic acid target, or its complementary sequence, and a copy of the label-tag; and

    c) detecting the plurality of amplified labeled-targets by sequencing at least a portion of the target and the label-tag; and

    d) determining the number of copies of the nucleic acid target, as indicated by the number of different label-tags that are associated with the nucleic acid target.

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