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METHODS AND COMPOSITIONS FOR ENRICHMENT OF TARGET POLYNUCLEOTIDES

  • US 20140162278A1
  • Filed: 12/10/2013
  • Published: 06/12/2014
  • Est. Priority Date: 07/17/2012
  • Status: Abandoned Application
First Claim
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1. A method of enriching a plurality of different target polynucleotides in a sample, the method comprising:

  • (a) joining an adapter oligonucleotide to each of the target polynucleotides, wherein the adapter oligonucleotide comprises sequence Y;

    (b) hybridizing a plurality of different oligonucleotide primers to adapted target polynucleotides, wherein each oligonucleotide primer comprises sequence Z and sequence W;

    wherein sequence Z is common among all oligonucleotide primers; and

    further wherein sequence W is different for each different oligonucleotide primer, is positioned at the 3′

    end of each oligonucleotide primer, and is complementary to a sequence comprising a causal genetic variant or a sequence within 200 nucleotides of a causal genetic variant;

    (c) in an extension reaction, extending the oligonucleotide primers along the adapted target polynucleotides to produce extended primers comprising sequence Z and sequence Y′

    , wherein sequence Y′

    is complementary to sequence Y; and

    (d) exponentially amplifying the purified extension products using a pair of amplification primers comprising (i) a first amplification primer comprising sequence V and sequence Z, wherein sequence Z is positioned at the 3′

    end of the first amplification primer; and

    (ii) a second amplification primer comprising sequence X and sequence Y, wherein sequence Y is positioned at the 3′

    end of the second amplification primer;

    wherein sequences W, Y, and Z are different sequences and comprise 5 or more nucleotides each.

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