Multiplex detection of nucleic acids

Multiplex detection of nucleic acids

  • CN 105,934,523 B
  • Filed: 11/26/2014
  • Issued: 12/29/2020
  • Est. Priority Date: 12/02/2013
  • Status: Active Grant
First Claim
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1. A method of quantifying a first chromosome or chromosomal locus relative to a second chromosome or chromosomal locus in a sample, comprisingContacting the sample with a first set of probes and a second set of probes, wherein the probes of the first set each specifically recognize a different target sequence within the first chromosome or chromosomal locus and wherein the probes of the second set each specifically recognize a different target sequence within the second chromosome or chromosomal locus,providing conditions such that the target sequence in the first and second chromosome or chromosomal locus becomes at least partially single-stranded,providing conditions for annealing and ligation under which the probe hybridizes to its target sequence and ligation products are produced, each ligation product being a nucleic acid loop comprising a ligation junction,providing conditions for rolling circle replication of the nucleic acid ring,labeling the rolling circle replication product by hybridizing the rolling circle replication product to a fluorescently labeled oligonucleotide to produce a fluorescently labeled rolling circle replication product,depositing the fluorescently labeled rolling circle replication product on the surface of the vector,counting the number of first rolling circle replication products on the vector, wherein the first rolling circle replication products are labeled with a first fluorophore and amplified from ligation products generated by the first set of probes to provide a first count,counting the number of second rolling circle replication products on the vector, wherein the second rolling circle replication products are labeled with a second fluorophore and amplified from ligation products generated by the second set of probes to provide a second count, andcomparing the first and second counts to determine the relative amounts of the first and second nucleic acid species in the sample,wherein the method is not used for diagnostic purposes.

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