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End-complementary polymerase reaction

  • US 6,489,146 B2
  • Filed: 02/05/1999
  • Issued: 12/03/2002
  • Est. Priority Date: 04/18/1995
  • Status: Expired due to Fees
First Claim
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1. A method of multiplex amplification, comprisingcontacting at least first and second noncontiguous polynucleotide sequences with at least first, second, third and fourth bivalent primers under amplification conditions, wherein the 3′

  • end of the first bivalent primer is complementary to the 3′

    end of the first polynucleotide sequence, and to the 5′

    end of the second bivalent primer, the 3′

    end of the second bivalent primer is complementary to the 3′

    end of the second polynucleotide sequence and to the 5′

    end of the first bivalent primer, the 3′

    end of the third bivalent primer is complementary to the 3′

    end of the complement of the first polynucleotide sequence, and to the 5′

    end of the fourth bivalent primer;

    the 3′

    end of the fourth bivalent primer is complementary to the 3′

    end of the complement of the second polynucleotide sequence, and to the 5′

    end of the third bivalent primer;

    the first, and second bivalent primers do not form stable hybrids with each other due to lack of sequence identity in internal segments of the first and second bivalent primers;

    the third and fourth bivalent primers do not form stable hybrids with each other due to lack of sequence identity in internal segments of the third and fourth bivalent primers;

    conducting a multi-cyclic amplification reaction to form a contiguous amplification product comprising equimolar amounts of the first and second polynucleotide sequences and an internal segment from one of the bivalent primers.

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