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Reactions on a solid surface

  • US 7,432,048 B2
  • Filed: 12/08/2000
  • Issued: 10/07/2008
  • Est. Priority Date: 11/29/1996
  • Status: Expired due to Fees
First Claim
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1. A method for cleaving multiple oligonucleotides comprising:

  • a) providing;

    i. a plurality of first oligonucleotides attached to a single solid support;

    ii. a second oligonucleotide attached to the same solid support as said plurality of first oligonucleotides;

    iii. a target nucleic acid, wherein said target nucleic acid comprises a first region and a second region, said second region downstream of and contiguous to said first region, wherein at least a portion of each of said first oligonucleotides is completely complementary to said first region of said target nucleic acid, and wherein said second oligonucleotide comprises a 3′

    portion and a 5′

    portion, wherein said 5′

    portion is completely complementary to said second region of said target nucleic acid; and

    iv. a cleavage agent comprising a FEN-1 endonuclease from an archaebacterial species;

    andb) exposing said solid support to said cleavage agent under conditions such that;

    i. a first invasive cleavage structure is formed, said first invasive cleavage structure comprising a first one of said plurality of first oligonucleotides, said second oligonucleotide, and said target nucleic acid;

    ii. said cleavage agent cleaves said first one of said plurality of first oligonucleotides in said first invasive cleavage structure to produce a first cleavage fragment;

    iii. after cleavage of said first one of said plurality of first oligonucleotides, a second invasive cleavage structure is formed, said second invasive cleavage structure comprising a second one of said plurality of first oligonucleotides and the same second oligonucleotide as was used in said first invasive cleavage structure;

    iv. said cleavage agent cleaves said first oligonucleotide in said second cleavage structure to produce a second cleavage fragment.

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