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HIGH THROUGHPUT DETECTION OF MICRORNAS AND USE FOR DISEASE DIAGNOSIS

  • US 20090181390A1
  • Filed: 10/31/2008
  • Published: 07/16/2009
  • Est. Priority Date: 01/11/2008
  • Status: Abandoned Application
First Claim
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1. A method for detecting a target nucleic acid in a sample comprisingproviding a sample potentially containing the target nucleic acidproviding at least one oligonucleotide probe set, wherein said probe set comprises:

  • (i) a first oligonucleotide probe having a 5′

    target specific region and a first 3′

    universal sequence region,(ii) a second oligonucleotide probe having a 3′

    target specific region and a second 5′

    universal sequence region(iii) a third oligonucleotide probe having a 5′

    region complementary to said first 3′

    universal sequence region in said first oligonucleotide probe, and(v) a fourth oligonucleotide probe having a 3′

    region complementary to the 5′

    universal sequence region of said second oligonucleotide probewherein the first and said second oligonucleotides probes are suitable for ligation together when hybridized adjacent to one another to said target nucleic acid, and wherein the third and the fourth oligonucleotides probes are suitable for ligation to the target nucleic acid when hybridized adjacent to said nucleic acid target;

    annealing the oligonucleotide probe set to the target nucleic acid such that a complex is formed between the target nucleic acid and the oligonucleotide probe set;

    contacting the complex with a linking agent under conditions such that the directly adjacent 5′ and

    3′

    ends of the first and second oligonucleotide probes, the 3′ and

    5′

    ends of the third oligonucleotide probe and the target nucleic acid, and the 5′ and

    3′

    ends of the fourth oligonucleotide probe and the target nucleic acid covalently bond to form a ligated probe product,separating the ligated probe product from the non-ligated first and second oligonucleotide probes; and

    detecting whether or not said ligated probe product is formed, wherein the presence of the ligated probe product is indicative of presence of said target nucleic acid in said sample.

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