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Hybridization probes and methods

  • US 10,030,268 B2
  • Filed: 11/11/2015
  • Issued: 07/24/2018
  • Est. Priority Date: 11/11/2014
  • Status: Expired due to Fees
First Claim
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1. A method of generating a probe to a nucleic acid target, comprising:

  • a) amplifying a plurality of regions of said nucleic acid target that are at least 85% free of undesired sequences wherein said undesired sequences are at least 100 bp in length to generate amplification products in a first amplification, wherein said amplifying comprises a primer pair complementary to said plurality of regions;

    b) combining said amplification products to generate a mixture;

    c) isolating said amplification products in said mixture;

    d) fragmenting said mixture of said isolated amplification products to generate a mixture of fragmented amplification products;

    e) size fractionating said fragmented amplification products;

    f) ligating said size fractionated amplification products to nucleic acid adaptors wherein said adaptors are each ligated to two or more amplification products, and wherein said adaptors comprise amplification primer sequences;

    g) amplifying said amplification products comprising said adaptors to generate amplification products in a second amplification wherein said amplifying comprises a primer pair complementary to said adaptor primer sequences; and

    h) labeling said amplification products generated in step g) to generate one or more said probes.

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